Significant Titers Research Articles

Loss of the vaccinia virus 35-amino acid hydrophobic O3 protein is partially compensated by mutations in the transmembrane domains of other entry proteins.

Eleven highly conserved proteins comprise the poxvirus entry-fusion complex (EFC). We focused on vaccinia virus (VACV) O3, a 35-amino acid, largely hydrophobic component of unknown specific function. Experimental evolution was carried out by blindly passaging a virus that was severely impaired in entry due to deletion of the gene encoding O3. Large plaque variants that arose spontaneously were discerned by round four and their numbers increased thereafter. Genome sequencing of individual cloned viruses revealed mutations in predicted transmembrane domains of three open reading frames encoding proteins with roles in entry. There were frame-shift mutations in consecutive Ts in open reading frames F9L and D8L and a nonsynonymous base substitution in L5R. F9 and L5 are EFC proteins and D8 is involved in VACV cell attachment. The F9L mutation occurred by round four in each of three independant passages, whereas the L5R and D8L mutations were detected only after nearly all of the genomes already had the F9L mutation. Viruses with deletions of O3L and single or double F9L, L5R and D8L mutations were constructed by homologous recombination. In a single round of infection, viruses with adaptive mutations including F9L alone or in combination exhibited statistically significant higher virus titers than the parental O3L deletion mutant or the L5R or D8L mutants, consistent with the order of selection during the passages. Further analyses indicated that the adaptive F9L mutants also had higher infectivities, entered cells more rapidly and increased EFC assembly, which partially compensated for the loss of O3.IMPORTANCE Entry into cells is an essential first step in virus replication and an important target of vaccine- elicited immunity. For enveloped viruses, this step involves the fusion of viral and host membranes to form a pore allowing entry of the genome and associated proteins. Poxviruses are unique in that this function is mediated by an entry-fusion complex (EFC) of eleven transmembrane proteins rather than by one or a few. The large number of proteins has hindered investigation of their individual roles. We focused on O3, a predominantly hydrophobic 35 amino acid component of the vaccinia virus EFC, and found that spontaneous mutations in the transmembrane domains of certain other entry proteins can partially compensate for the absence of O3. The mutants exhibited increased infectivity, entry and assembly or stability of the EFC.

Increasing the efficiency of diagnostics of periodontal tissue diseases with modern research method

Nowadays periodontal tissue diseases remain the leading dental pathology. Healthy periodontium is revealed only in 12% of people, while the rest have lesions of varying severity: from initial inflammatory to severe destructive changes with loss of teeth. Severe periodontitis, leading to the loss of teeth, is found in 5-15% of cases. To diagnose and prescribe treatment, laboratory assessment of the content of the gingival groove and / or periodontal pocket was studied. We assessed the titer of the aerobic and anaerobic microorganisms in the samples, the greatr amount was in a clinically significant titer. Although there are various instruments for taking the contents of the gingival groove and / or periodontal pocket, they have a number of drawbacks which complicate the culturing of the material and distort the findings. Taking into account the advantages and disadvantages of the existing instruments, we have developed the instrument for collecting the content of the gingival groove.

An organoid‐derived bronchioalveolar model for SARS‐CoV‐2 infection of human alveolar type II‐like cells

Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) causes coronavirus disease 2019 (COVID‐19), which may result in acute respiratory distress syndrome (ARDS), multiorgan failure, and death. The alveolar epithelium is a major target of the virus, but representative models to study virus host interactions in more detail are currently lacking. Here, we describe a human 2D air–liquid interface culture system which was characterized by confocal and electron microscopy and single‐cell mRNA expression analysis. In this model, alveolar cells, but also basal cells and rare neuroendocrine cells, are grown from 3D self‐renewing fetal lung bud tip organoids. These cultures were readily infected by SARS‐CoV‐2 with mainly surfactant protein C‐positive alveolar type II‐like cells being targeted. Consequently, significant viral titers were detected and mRNA expression analysis revealed induction of type I/III interferon response program. Treatment of these cultures with a low dose of interferon lambda 1 reduced viral replication. Hence, these cultures represent an experimental model for SARS‐CoV‐2 infection and can be applied for drug screens.

Immunogenicity and protective efficacy of BBV152, whole virion inactivated SARS- CoV-2 vaccine candidates in the Syrian hamster model

SummaryThe availability of a safe and effective vaccine would be the eventual measure to deal with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) threat. Here, we have assessed the immunogenicity and protective efficacy of inactivated SARS-CoV-2 vaccine candidates BBV152A, BBV152B, and BBV152C in Syrian hamsters. Three dose vaccination regimes with vaccine candidates induced significant titers of SARS-CoV-2-specific IgG and neutralizing antibodies. BBV152A and BBV152B vaccine candidates remarkably generated a quick and robust immune response. Post-SARS-CoV-2 infection, vaccinated hamsters did not show any histopathological changes in the lungs. The protection of the hamster was evident by the rapid clearance of the virus from lower respiratory tract, reduced virus load in upper respiratory tract, absence of lung pathology, and robust humoral immune response. These findings confirm the immunogenic potential of the vaccine candidates and further protection of hamsters challenged with SARS-CoV-2. Of the three candidates, BBV152A showed the better response.

Concurrent Anti–Glomerular Basement Membrane Antibody Disease and Membranous Nephropathy: A Case Series

Anti-glomerular basement membrane (GBM) disease is a rapidly progressive glomerulonephritis which, in some instances, occurs concurrently with other diseases such as antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis. Rarely, it also occurs with membranous nephropathy (MN). We report a series of such patients, characterizing their long-term follow up. Case series. Twelve patients referred to 1 medical center between 2001 and 2019 with anti-GBM disease and MN whose clinical characteristics and course were described. Patients' ages ranged from 20 to 81 years old, and all presented with severe acute kidney injury requiring dialysis on presentation or shortly thereafter. Only 1 patient had pulmonary findings on presentation. The predominant pathology was crescentic and necrotizing glomerulonephritis with linear staining for immunoglobulin G along the GBM associated with epimembranous electron-dense immune-type deposits. All 11 patients who were tested had significant titers of anti-GBM antibodies, but only 1 of the 5 tested for phospholipase A2 receptor (PLA2R) on biopsy was positive. Eight patients received therapy with cyclophosphamide, prednisone, and plasmapheresis; 2 patients with prednisone and plasmapheresis; and 2 with rituximab-based regimens. Progression to a requirement for kidney replacement therapy occurred in all 12 patients, but 2 patients later recovered kidney function. Recurrence of anti-GBM disease did not occur for any of the patients studied. Incomplete testing for PLA2R in biopsy and serum, limited sample size, and lack of uniform treatment regimen. In this case series, the presentation of concurrent anti-GBM disease and MN was characterized by rapidly progressive glomerulonephritis and poor kidney outcomes. These findings suggest possible value from earlier diagnosis and the need for identification of more effective treatment regimens.

A cross-sectional descriptive study of clinical and serological prevalence of syphilis infection in people living with HIV and its effect on CD4+ T cells.

Context:The natural history of syphilis could be altered in the presence of HIV. It has been documented that syphilis infection increases the risk of HIV transmission by at least 3-fold.Aims:The aim of the study was (1) to study clinical presentation of syphilis in HIV individuals, (2) to estimate seroprevalence of syphilis in HIV individuals, and (3) to study the effect of syphilis infection on CD4+ T cells.Subjects and Methods:HIV-positive patients attending sexually transmitted infection clinic for a period of 1 year from June 2017 to May 2018 in the age group of 15–70 years were included in the study. A detailed history was taken; genital and dermatological examination was done. All patients were tested with VDRL and treponema pallidum hemagglutination assay (TPHA). Pearson's Chi-square test was used to compare categorical variables and Student's t-test was used to compare continuous variables.Results:Out of ninety study population, nine (10%) had clinical manifestations of syphilis. VDRL was positive with significant titers in all cases of syphilis. TPHA was positive in 88.9% of cases with clinical syphilis and 17.3% of cases without clinical manifestations of syphilis. Mean CD4 count was low among patients having syphilis infection compared to study population.Conclusion:This study shows high prevalence of syphilis in HIV and highlights the importance of preventing and promptly treating syphilis in people living with HIV, as the active infection is associated with fall in CD4 count, which leads to opportunistic infections.

Monitoring of SARS-COV-2 antibodies in tuberculosis in-patient clinic stuff

Objective. Тo study the features of humoral immunity to SARS-CoV-2 among the medical stuff of an antitubercular hospital in the context of the spread of a new coronavirus infection. Materials and methods. The interim results of a single-center prospective observational study involving 350 employees of a pulmonary tuberculosis hospital that was not repurposed to work with COVID-19 are presented. The results of weekly monitoring of IgM and IgG levels for the period from May 15 to November 28, 2020, were analyzed in comparison with the results of diagnosis of COVID-19 cases (identification of the pathogen in a smear from the oropharynx and nasopharynx, changes in lung tissue according to computed tomography). Results. During the observation period, COVID-19 was detected in 106 people (30.3%). Antibodies to SARS-CoV-2 (IgG and/or IgM) were detected at the start of monitoring in 30 people (8.6%, 95%CI 5.8-12.2%), by the end of the follow – up period-in 95 people (27.1%, 95%CI 22.7- 32.0%). The frequency of detection of antibodies in the patients was 70.8% (95%CI 55.6-88.7%): IgM class in 23 (21.7%), G class in 73 (68.9%). The median time from disease detection to IgM detection was 1.5 weeks. (interquartile range 0-4 weeks), before the discovery of IgG – 4 weeks. (interquartile range 2-5 weeks). At any follow-up period, the chances of IgG production in the protective titer were higher in the case of moderate or severe disease with the development of pneumonia compared to the mild course (OR 2.6, 95%CI 1.1-6.3); the IgG titer was also higher (p < 0.01). «Atypical» variants of the antibody response were identified in individuals with confirmed COVID-19: no antibody formation (31 people), production of class M antibodies only (4 people); long-term persistence of IgG in a titer of less than 30 g/l, long ahead of the disease manifestation (6 people). The sensitivity of serological testing as a method for detecting current and / or transmitted coronavirus infection was 70.8% (95%CI 61.1-79.2%), and the specificity was 91.8% (95%CI 87.6-94.9%). Conclusion. The frequency of detection of antibodies to SARS-CoV-2 among employees of the TB hospital corresponds to the dynamics of the incidence of COVID-19. Virus-specific antibodies were detected in 70.8% of COVID-19 survivors (IgG in 68.9%); in the absolute majority of patients, the antibodies remained in a significant titer (IgG > 10 g/l) for 6 months or more at the start of testing. When the disease occurs without the development of viral pneumonia, the chances of forming a significant and persistent antibody response are significantly lower. Serological testing is a sensitive and highly specific method for detecting a previous COVID-19 disease.

Retrospective serological study of Rickettsia spp. and Borrelia spp. antibodies in patients with peripheral facial nerve palsy

ABSTRACT In a retrospective study, 36 patients with peripheral facial palsy were serologically evaluated for the presence of Rickettsia spp. and Borrelia spp. antibodies. All sera underwent immunofluorescence and Western blot analysis for IgG and IgM antibodies using Rickettsia helvetica and R. felis as antigens. Anti-Borrelia antibodies were detected using a commercial ELISA detecting Borrelia burgdorferi, B. afzelii and B. garinii. Three patients (8.3%) were seropositive for Rickettsia spp. with IgG titres equal to 1:128, and six patients (16.7%) had IgM titres equal to or above 1:128. All samples with IgG/IgM titres equal to or above 1:128 were confirmed by Western Blot. Four patients (11.1%) had IgG antibodies against Borrelia at a titre level normally judged to be indicative of current infection. Two of these patients had significant IgG or IgM titres for both Rickettsia spp. and Borrelia spp., indicating co-infection. In conclusion, the findings indicate current rickettsial infection or early response at about the same degree as for Lyme borreliosis in patients with facial palsy, but they need to be further examined with a larger number of patients and paired serum analyses.

Recombinant outer membrane protein OmpC induces protective immunity against Aeromonashydrophila infection in Labeorohita

Aeromonashydrophila is an opportunistic pathogen that causes enormous loss to aquaculture industry. The outer membrane proteins of Aeromonas help in bacterium-host interaction, and are considered to be potential vaccine candidates. In the present study, we evaluated immunogenicity and protective efficacy of recombinant OmpC (rOmpC) of A. hydrophila in Indian major carp, Labeorohita. The rOmpC-vaccinated fish produced specific anti-rOmpC antibodies with a significant antibody titer, and the antisera could specifically detect the rOmpC in the cell lysates of Escherichia coli expressing rOmpC and cross-react with different Aeromonas lysates, indicating the suitability of the anti-rOmpC antisera to detect Aeromonas infection. A significant increase was noted in ceruloplasmin level, myeloperoxidase and anti-protease activities in transient and temporal manner the sera of the rOmpC-immunized fish as compared to PBS-control fish. Higher agglutination- and hemolytic activity titers in the anti-rOmpC antisera indicate stimulation of innate immunity. Expression of immune-related genes comprising various acute phase proteins, cytokines and inflammatory response molecules were modulated in the head kidney of rOmpC-immunized L. rohita. While IgM, IL1β, and TLR-22 were significantly up-regulated at early time points (3 h–72 h), the others showed a transient augmentation at both early and later time points (SOD, lysozymes C and G, NKEF-B, C3, CXCa and TNF-α) in the rOmpC-immunized L. rohita in comparison to PBS-injected controls. These data suggest that the rOmpC-induced immune response is temporally regulated to confer immunity. In vivo challenge of the rOmpC-immunized fish with A. hydrophila showed significantly greater survival when compared to PBS-injected control fish. Thus, our results highlight the immunomodulatory role of rOmpC and demonstrate its protective efficacy in L. rohita, along with the use of anti-rOmpC antisera in detecting Aeromonas infections.

Assessment of Mouse Ileal loop Protection against Clinically Isolated Vibrio cholerae Outer Membrane Vesicles as a Vaccine Candidate

Cholera, a life-threatening disease caused by the Gram-negative bacterium Vibrio cholera, remains a concern in developing countries. The present study investigated the immunogenicity and protective immunity of outer membrane vesicles (OMVs) and combination of OMV and killed whole cells (WC) of a local strain isolated from the last outbreak in Iran in addition to reference and local strains of V. cholerae El Tor O1 in comparison to Dukoral vaccine in mice model. The protein content, morphology, and size of extracted OMVs were evaluated by electrophoresis and microscopic analyses, respectively. The serum titers of total immunoglobulin G (IgG), IgG1, IgG2a, and immunoglobulin A (IgA) in addition to secretory IgA and total IgG in different mice groups were determined by enzyme-linked immunosorbent assay (ELISA). In addition, fluid accumulation (FA) assay regarding the resistance to live strain of V. cholerae in ligated ileal loops was carried out to determine immunogenicity by OMV or combination of OMV and WC in comparison to that reported for Dukoral vaccine. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of purified OMVs indicated protein profiles within the range of 34-52 kDa. Furthermore, transmission electron microscopy demonstrated the spherical shaped vesicles of 50-200 nm. The results of ELISA showed significant titers of systemic and mucosal immune anti-OMV IgGs in immunized BALB/c mice with different vaccine regimens. Additionally, a notable increase in the FA ratio was demonstrated in this study. The obtained results of the present study revealed that the WC-OMV combination of local strain can induce a high level of antibody response indicating more protection than OMV or WC separately. Moreover, it can be considered an effective immunogen against V. cholerae.

Oral immunization of rat with chromosomal expression LipL32 in attenuated Salmonella vaccine induces immune respond against pathogenic Leptospira.

PurposeLeptospirosis caused by Leptospira spp. remains a global health problem. Available commercial leptospiral vaccines have shown an ineffective prevention for leptospiral infection. The aim of this study was to develop leptospirosis vaccine using recombinant attenuated Salmonella vaccine (RASV) as a platform. We expected that this vaccine has ability to continuous and strongly stimulate immune systems including protective mucosal, humoral, and cell mediated immunity in rat model.Materials and MethodsIn this study, we engineered RASV, NRSL32 strain containing chromosomal fusion between nucleotides encoding secretion signal of SPI-2 effector protein, SspH2 and gene encoding major pathogenic leptospiral outer membrane lipoprotein, LipL32. Subsequently, our modified RASV was oral vaccination to rat and blood samples were taken for assessment of immune responses.ResultsOur Salmonella NRSL32 strain showed expression and secretion of SspH21-215-LipL32 recombinant protein via SPI-2 T3SS. After oral administration of NRSL32 strain to rats, significant titers of total immunoglobulin G (IgG) and immunoglobulin A against rLipL32 were observed in long period up to 77 days after vaccination. The stimulated antibody showed ability to specific bind with LipL32 protein on surface of pathogenic Leptospira spp. Additionally, the balance level of IgG2a/IgG1 ratio and level of interferon-γ and interleukin-4 secretion were detected.ConclusionThe results showed that our RASV platform with chromosomal expression elicited effective immune responses to leptospiral antigen. Moreover, this platform was capable for simultaneous stimulation of Th1 and Th2-biased responses. Further investigation is necessary study of protective efficacy against leptospiral infection in animal models.

Guillain-Barré syndrome during the Zika virus outbreak in Northeast Brazil: An observational cohort study

ObjectiveTo determine the clinical phenotype of Guillain-Barré syndrome (GBS) after Zika virus (ZIKV) infection, the anti-glycolipid antibody signature, and the role of other circulating arthropod-borne viruses, we describe a cohort of GBS patients identified during ZIKV and chikungunya virus (CHIKV) outbreaks in Northeast Brazil. MethodsWe prospectively recruited GBS patients from a regional neurology center in Northeast Brazil between December 2014 and February 2017. Serum and CSF were tested for ZIKV, CHIKV, and dengue virus (DENV), by RT-PCR and antibodies, and serum was tested for GBS-associated antibodies to glycolipids. ResultsSeventy-one patients were identified. Forty-eight (68%) had laboratory evidence of a recent arbovirus infection; 25 (52%) ZIKV, 8 (17%) CHIKV, 1 (2%) DENV, and 14 (29%) ZIKV and CHIKV. Most patients with a recent arbovirus infection had motor and sensory symptoms (72%), a demyelinating electrophysiological subtype (67%) and a facial palsy (58%). Patients with a recent infection with ZIKV and CHIKV had a longer hospital admission and more frequent mechanical ventilation compared to the other patients. No specific anti-glycolipid antibody signature was identified in association with arbovirus infection, although significant antibody titres to GM1, GalC, LM1, and GalNAc-GD1a were found infrequently. ConclusionA large proportion of cases had laboratory evidence of a recent infection with ZIKV or CHIKV, and recent infection with both viruses was found in almost one third of patients. Most patients with a recent arbovirus infection had a sensorimotor, demyelinating GBS. We did not find a specific anti-glycolipid antibody signature in association with arbovirus-related GBS.

Phase I Trial of a Ligand Fusion-Protein (DAB389IL-2) in Lymphomas Expressing the Receptor for Interkeukin-2

AbstractThe purpose of this study was to evaluate the safety, tolerability, pharmacokinetics, and possible antitumor activity of a ligand fusion-protein, DAB389IL-2, in a phase I trial. This was a multicenter, open-label, dose-escalation trial. Patients with preserved organ function and histologically confirmed relapsed cutaneous T-cell lymphoma (CTCL), other non-Hodgkin's lymphomas (NHL), or Hodgkin's disease (HD) were eligible if their cancer was shown to express the interleukin (IL)-2 receptor by an immunohistochemical assay for the p55 or the p75 subunit. Patients received up to eight courses of DAB389IL-2 given as a short intravenous infusion daily for 5 days with subsequent courses every 21 days. The maximum tolerated dose (MTD) and tumor response was determined according to standard criteria. Seventy-three patients (44 men/29 women), aged 16 to 81 years (mean, 50.7) with CTCL (n = 35), NHL (n = 17), and HD (n = 21) were enrolled. The patients were extensively treated, failing 0 to 15 previous therapies (median, 4). Patients received one to six courses (mean, 3.3) of DAB389IL-2 over a range of 3 to 31 μg/kg/day. The dose-limiting toxicity was asthenia, establishing the maximum tolerated dose of 27 μg/kg/day. Approximately half of all patients had significant titers of antibody to diphtheria toxin or to DAB389IL-2 at the time of enrollment compared with 92% with titers at the end of treatment. The presence of antibody did not preclude clinical response. There were five complete (CR) and eight partial (PR) remissions in patients with CTCL with one CR and two PR occurring in NHL. The median time to response was 2 months and the duration of response was 2 to 39+ months. No responses were documented in patients with HD. DAB389IL-2 is well tolerated with an MTD of 27 μg/kg/day. This ligand fusion-protein showed antitumor effects in patients with IL-2 receptor expressing CTCL and NHL. Additional trials in these diseases are warranted.

Abo不相容性和抗a / B异凝集素滴度对血液恶性肿瘤清髓处理后输血需求和无关脐带血移植的早期结果的影响

Abstract Background:ABO incompatibility is not considered a main contraindication to allogeneic hematopoietic stem cell transplantation (aHSCT). However, it has been associated with a number of immunohematological complications. The effects of ABO incompatibility on aHSCT remain controversial.The change of isoagglutinin titers and early clinical outcomes were analyzed after unrelated cord blood transplantation (UCBT) with ABO-incompatibility donor. Methods:252 patients with hematological malignant diseases and other hematological disorders who underwent unrelated UCBT from January 2019 to April 2020 were retrospectively analyzed in this research. Patients were studied in identical, major, minor and bidirectional mismatch groups. Immunoglobulin m (IgM) isoagglutinin titers were tested one day before the transplant (-1 day), 2 weeks post-transplant, 4 weeks post-transplant and 6 weeks post-transplant. R esults:76 match,71 major mismatch, 70 minor mismatch and 35 bidirectional unrelated UCBT were identified. The median neutrophil, PLT and red blood cell (RBC) recovery days were 18, 38 and 22, respectively. ABO mismatch did not influence the neutrophil, PLT and RBC engraftment. The median of RBC transfusion in 30 days were 5 units and PLT were 6 units. There were no statisitcal difference in 0-30 days RBC and PLT transfusion after UCBT. 31-100 days transfusion was similar to in 30 days transfusion. No patients developed pure red cell anemia (PRCA). -1day IgM titers ≥1:16 did not develop higher risk of grade II-IV aGVHD when compared with titers≤1:8 group. However, we detected a marginal higher PLT transfusion in 30 days after transplant at antibody titers ≥1:16 group when compared with titers≤1:8 (P=0.051). In the major and bidirectional groups, we found that group O IgM anti-donor antibodies were displayed a significant higher than the group B anti-A titer (p<0.001) in setting the time one day before the transplant, but no significant with group A. 2 weeks after the transplant, group B anti-A was still showed significant lower than the group O anti-A (p<0.001). 4 weeks after the UCBT, we observed a modest, but no statistical significant lower titers of group B anti-A antibodies as compared with O group (P=0.097). 6 weeks after the transplant, there were no statistical significant among group O, A and B. In the multivariable Cox regression model, transfusion of ≥5 RBC units in 30 days after UCBT (HR=1.727, 95%CI=1.020-2.926, P=0.042) and PLT engraftment ≥38 days (HR=1.964, 95%CI=1.134-3.401, P=0.016) were correlated to greater risk of grade severe aGVHD. Conclusion:This study showed that ABO mismatch did not influence the neutrophil, PLT and RBC recovery time.Group O IgM anti-donor isoagglutinins in recepients showed a higher titers than the group B in setting with the time (-1 days pre-transplant, 2 weeks post-transplant, 4 weeks post-transpant). Pre-transplant higher anti-donor isoagglutinins were associated with more PLT transfusion requirements after UCBT. More RBC transfusion (≥5 units) and longer PLT recovery time (≥38 days) showed a higher incidence of severe aGVHD. Disclosures No relevant conflicts of interest to declare.

Optimized Adenoviral Vector That Enhances the Assembly of FMDV O1 Virus-Like Particles in situ Increases Its Potential as Vaccine for Serotype O Viruses.

Although replication-defective human adenovirus type 5 (Ad5) vectors that express in situ the capsid-encoding region of foot-and-mouth disease virus (FMDV) have been proven to be effective as vaccines in relevant species for several viral strains, the same result was not consistently achieved for the O1/Campos/Brazil/58 strain. In the present study, an optimization of the Ad5 system was explored and was proven to enhance the expression of FMDV capsid proteins and their association into virus-like particles (VLPs). Particularly, we engineered a novel Ad5 vector (Ad5[PVP2]OP) which harbors the foreign transcription unit in a leftward orientation relative to the Ad5 genome, and drives the expression of the FMDV sequences from an optimized cytomegalovirus (CMV) enhancer-promoter as well. The Ad5[PVP2]OP vaccine candidate also contains the amino acid substitutions S93F/Y98F in the VP2 protein coding sequence, predicted to stabilize FMD virus particles. Cells infected with the optimized vector showed an ∼14-fold increase in protein expression as compared to cells infected with an unmodified Ad5 vector tested in previous works. Furthermore, amino acid substitutions in VP2 protein allowed the assembly of FMDV O1/Campos/Brazil/58 VLPs. Evaluation of several serological parameters in inoculated mice with the optimized Ad5[PVP2]OP candidate revealed an enhanced vaccine performance, characterized by significant higher titers of neutralizing antibodies, as compared to our previous unmodified Ad5 vector. Moreover, 94% of the mice vaccinated with the Ad5[PVP2]OP candidate were protected from homologous challenge. These results indicate that both the optimized protein expression and the stabilization of the in situ generated VLPs improved the performance of Ad5-vectored vaccines against the FMDV O1/Campos/Brazil/58 strain and open optimistic expectations to be tested in target animals.

Molecular associations of gallinacin genes with immune response against Salmonella typhimurium in chickens

Candidate gallinacin genes (GAL) were assessed in Fayoumi (F), Rhode Island Red (R) and their crosses (½R½F and ½F½R) using PCR-RFLP technique to detect the associations between GAL 2, GAL 3, GAL 4 and GAL 5 genes and caecal S. typhimurium bacterial count (CSTBC) and IgA, IgY and IgM antibody titers. The solutions of genotypes of GAL genes were calculated by the method of Generalized Least Squares (GLS). The SNPs genotypes of GAL 3 and GAL 5 genes showed significant counts of caecal S. typhimurium. The SNP of gallinacin 3, 4 and 5 genes had significant effects on IgA, IgY and IgM antibody titers. For GAL 3 gene, the chicks of genotype CC in R breed had lower significant CSTBC and higher significant IgA and IgM antibody titers than chicks of TT genotype, while the chicks of TC genotype had lower significant CSTBC in chicks of ½R½F crossbred and higher significant antibody titers of IgA and IgM in chicks of ½F½R crossbred. For GAL 4 gene, the chicks of genotype GG in R breed had lower significant CSTBC and higher significant IgA, IgY and IgM antibody titers, but the chicks of genotype AG had higher significant IgA, IgY and IgM antibody titers in chicks of ½R½F crossbred than chicks of GG and AA genotypes. For GAL 5 gene, the genotype CC in chicks of R breed had lower significant CSTBC and higher significant IgA and IgY antibody titers. In chicks of ½F½R crossbred, the chicks of genotype AA had lower significant CSTBC and higher significant IgA and IgY antibody titers than chicks of CA genotype. In practice, GAL genes could be used as markers assisted selection to improve immune response against S. typhimurium in genetic improvement programs of chickens.

Look Beyond Gluten in Short Stature with Celiac Disease - A Prospective, Interventional Study.

Short stature (SS) is a common manifestation of celiac disease (CD). After starting gluten free diet (GFD), children usually have catch-up growth (improvement in height SDS of >1 SD). However, few children in remission, even on GFD, lack catch up growth. This study was planned to assess the growth hormone (GH) axis and the prevalence of anti-pituitary antibodies in such patients. It was a single-centre, prospective study. Patients with CD in remission for the last 1 y, having SS and lacking catch-up growth, were included after excluding other common causes of SS. GH dynamics were studied using stimulation tests: Insulin tolerance test, clonidine stimulation test, and glucagon stimulation test. GH deficiency (GHD) was defined as non-stimulable response to 2 GH stimulation tests. Anti-pituitary antibodies were analysed in these patients using rat pituitary extract as antigen. Ten patients (8 girls), with a mean age of 10 ± 2.8 y, in serological remission for CD and lacking catch-up growth, were enrolled. All had a height SDS of < -2. Fifteen age matched children with CD and adequate catch up growth served as controls. GHD was seen in 7 patients (70%), out of whom 2 received GH therapy and had an improvement in the height SDS from -2.7 to -1.4 and from -2.1 to 2.4 (over 1 y), respectively. Anti-pituitary antibodies were seen in significant titres in 55.5% of the cases and 40% of the controls. Children with CD in remission but lacking catch-up growth should be evaluated for GHD.

The experience of using a bacteriophages-based complex antibacterial and analgesic drug in gel formulation in women who underwent various instrumental and diagnostic and treatment interventions

During the spread of bacterial infections with extensive drug resistance, the potential of existing antibacterial drugs is significantly reduced. The medical community is seeking opportunities for the rational use of antibacterial drugs, the development of new drugs, as well as the use of alternative treatments for these infections.&#x0D; Aim. The objective of this study was to evaluate the effectiveness of the use of a complex antibacterial and analgesic drug based on bacteriophages in gel form during various instrumental studies and diagnostic procedures in order to minimize the risk of the urinary system infections complications.&#x0D; Materials and methods. The study included 235 women aged 18 to 75 years who underwent treatment at the Moscow State Clinical Hospital named after Spasokukotsky and in the clinic of urology of Moscow State University of Medicine and Dentistry from September 2019 to January 2020. During the manipulation, all patients of the main group (n=120) used a developed preparation based on bacteriophages, and in the control group (n=115), a topical gel based on chlorhexidine dihydrochloride and lidocaine hydrochloride was used. All patients before the manipulation, as well as 10 days after the manipulation, underwent bacteriological analysis of urine. Efficiency assessment was carried out on the basis of laboratory data obtained, as well as according to the visual analogue scale of the pain syndrome severity.&#x0D; Results. None of the patients in the main group showed intolerance, side effects and allergic reactions of the developed drug based on bacteriophages. Bacteriological analysis of urine received before manipulation in both groups showed the absence of clinically significant titers of pathogenic bacteria, while 1 week after manipulation the number of titers of pathogenic bacteria (104) was significantly higher in the control group (p0.04). In patients of the main group, the incidence of dysuria was lower compared with the control group (p0.05).&#x0D; Conclusion. The study showed good tolerance of the developed drug and its clinical effectiveness in reducing the number of symptomatic bacteriuria; severity of pain after undergoing manipulation was relatively lower in the main group of patients. Conducting multicenter studies with the inclusion of a larger number of patients in the future will reduce the economic costs associated with treating patients by reducing the number of cases of nosocomial infections and reducing postoperative time spent by patients in the hospital.

Antimicrobial Activity of Chitosan-Based Films Enriched with Green Tea Extracts on Murine Norovirus, Escherichia coli, and Listeria innocua.

Edible films can be designed to serve as carriers of antimicrobial agents and be used to control pathogenic foodborne viruses and bacteria. This research tested this concept by dissolving green tea extract (GTE) in chitosan film-forming solutions (FFS) and using it to prepare dried chitosan edible films. As a control, the GTE was also dissolved in deionized water (DW). The FFS and the dried chitosan films with the GTE and the DW without chitosan were all evaluated against murine norovirus (MNV-1), Escherichia coli K12, and Listeria innocua. Both the FFS and the DW with GTE were incubated with ~107 PFU/ml of the virus suspensions for 3 h. The chitosan films with GTE were incubated for 4 and 24 h at 23 ± 1°C. The results showed that the DW containing 1, 1.5, and 2.5% aqueous GTE, significantly (p < 0.05) reduced MNV-1 plaques by 1.7, 2.5, and 3.3 logs after 3 h exposure, respectively. Similarly, FFS containing 2.5 and 5.0% GTE reduced MNV-1 counts by 2.5 and 4.0 logs, respectively, after 3 h exposure. The dried chitosan films with 5, 10, and 15% GTE were also effective against MNV-1 infectivity. After 24 h incubation, the 5 and 10% chitosan GTE films produced significant (p < 0.05) titer reductions of 1.6 and 4.5 logs, respectively. Chitosan films containing 15% GTE reduced MNV-1 plaques to undetectable levels in 24 h. All chitosan GTE films reduced E. coli K12 and L. innocua populations to undetectable levels in tryptic soy broth after 24 h exposure. The results of this study showed that edible films enriched with GTE have potential to reduce both foodborne viruses and bacteria.

Vaccination with dendritic cells pulsed ex vivo with gp100 peptide-decorated liposomes enhances the efficacy of anti PD-1 therapy in a mouse model of melanoma

BackgroundTargeting antigens to dendritic cells (DCs) via nanoparticles is a powerful strategy which improves the efficacy of ex vivo antigen-pulsed DC vaccines. MethodsIn this study, liposomes were first decorated with gp10025-33 self-antigen and then characterized. Then, DCs were pulsed ex vivo with liposomal gp100 and injected subcutaneously in mice bearing B16F10 established melanoma tumors in combination with anti-PD-1 therapy. ResultsTreatment with liposomal pulsed DC vaccine elicited the strongest anticancer immunity and enhanced intratumoral immune responses based on infiltration of gp100-specific CD4+ and CD8+ T cells to the tumor leading to significant tumor growth regression and prolonged survival rate. Treatment with liposomal pulsed DC vaccine also markedly enhanced specific cytotoxic T lymphocytes (CTL) responses with a significant higher titer of IFN-γ in the spleen. Moreover, a significant increase of PD-1 expressing CD8+ tumor infiltrating lymphocytes (TILs) was detected in tumors. ConclusionOur results demonstrate an optimum dose of liposomal gp100 significantly increases the efficacy of anti-PD-1 therapy in mice and might be an effective strategy to overcome resistance to anti-PD-1 therapy.