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Objetivo. Evaluar la respuesta serológica de anticuerpos de una llama (Lama glama) a la inmunización del virus SARS-CoV-2 (linaje B.1.1) y la capacidad neutralizante del suero de llama hiperinmune frente al virus SARS-CoV-2 (linaje B.1.1) en células Vero. Materiales y métodos. Se inmunizó una llama con el virus SARS-CoV-2 inactivado (Linaje B.1.1) y se analizaron muestras de suero para evaluar el nivel de anticuerpos mediante ELISA, así como la reactividad a antígenos de SARS-CoV-2 mediante Western Blot. Además, se evaluó la neutralización viral en cultivos celulares por la Prueba de Neutralización por Reducción de Placas (PRNT, por sus siglas en inglés). Resultados. Se observó un aumento en la serorreactividad en la llama inmunizada desde la semana 4 en adelante. Los títulos de anticuerpos fueron más elevados en el séptimo refuerzo de inmunización. Los resultados de Western Blot confirmaron los hallazgos positivos del ELISA, y los anticuerpos del suero inmune reconocieron varias proteínas virales. El ensayo de neutralización (PRNT) mostró una neutralización viral visible, concordante con los resultados de ELISA y Western Blot. Conclusiones. Los hallazgos sugieren que el suero hiperinmune de llama podría constituir una fuente de anticuerpos terapéuticos contra las infecciones por el virus SARS-CoV-2 (linaje B.1.1) y que deberá ser evaluado en estudios posteriores.
Objective. To evaluate the serological antibody response of a llama (Lama glama) to SARS-CoV-2 (B.1.1 lineage) immunization and the neutralizing capacity of hyperimmune llama serum against SARS-CoV-2 virus (B.1.1 lineage) in Vero cells. Materials and methods. A llama was immunized with inactivated SARS-CoV-2 (B.1.1 lineage). Serum samples were analyzed to evaluate the level of antibodies by ELISA, as well as reactivity to SARS-CoV-2 antigens by Western Blot. In addition, viral neutralization in cell cultures was assessed by the Plate Reduction Neutralization Test (PRNT). Results . Seroreactivity increased in the immunized llama from week 4 onwards. Antibody titers were the highest after the seventh immunization booster. Western blot results confirmed the positive ELISA findings, and immune serum antibodies recognized several viral proteins. The neutralization assay (PRNT) showed visible viral neutralization, which was in accordance with the ELISA and Western Blot results. Conclusions. The findings suggest that hyperimmune llama serum could constitute a source of therapeutic antibodies against SARS-CoV-2 infections (lineage B.1.1), and should be studied in further research.
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AnimalsABSTRACT
Objetivo: Optimizar la prueba de microneutralización (MNT) para detección de anticuerpos neutralizantes contra el virus dengue serotipo 2 (DENV-2) en la línea celular Vero-76. Materiales y métodos: Se evaluaron diferentes concentraciones celulares (0.6 x105 cel/mL, 0.9 x105 cel/mL, 1.2 x105 cel/mL), porcentajes de 2 %, 3 % y 4 % de suero bovino fetal (SBF), número de pasajes del stock de virus y los días de incubación. La semilla viral se confirmó por RT-qPCR. El DENV-2 se propagó realizando 5 pasajes en células Vero-76, seguidamente se tituló el virus en placas de 96 pozos y se evaluaron 2 métodos de infección celular: monocapa y células en suspensión, además se determinó el día óptimo de coloración de las células. Obtenidos los resultados, se procesaron mediante MNT para DENV-2 las siguientes muestras: 5 sueros negativos a DENV-2 y YFV (Virus de la Fiebre Amarilla), 5 sueros negativos de anticuerpos a DENV-2 y positivos de anticuerpos a YFV y 5 sueros positivos de anticuerpos a DENV-2 seleccionados mediante la prueba de neutralización por reducción de placas (PRNT). Resultados: El método óptimo para MNT utilizó células en suspensión (0.9 x 105 cel/mL), 2 % de SBF y semilla viral pasaje 5. La mínima dilución capaz de diferenciar una muestra positiva a DENV-2 fue 1:40 y el tiempo de incubación para la MNT para DENV-2 fue de 10 días. Conclusión: La MNT con el método de células en suspensión y medio de cultivo con 2 % de SBF permite detectar anticuerpos neutralizantes IgG contra DENV-2 con resultados confiables, pudiendo analizar un mayor número de muestras con ahorro de materiales.
Objective: To optimize the microneutralization test (MNT) for the detection of neutralizing antibodies against serotype 2 of dengue fever virus (DENV-2) in a Vero-76 cell line. Materials and methods: Different cell concentrations were assessed (0.6 X 105 cells/mL, 0.9 X 105 cells/mL, and 1.2 x 105 cells/mL) with 2%, 3%, and 4% fetal bovine serum, virus stock passage number, and incubation days. Viral particles were confirmed using RT-qPCR. DENV-2 disseminated with 5 passages in Vero-76 cells, then, the virus was titrated in 96-well plaques, and two methods for cell infection were evaluated: single layer, and suspended cells. Also, the optimum day for cell staining was determined. Once results were obtained, the following samples were processed using MNT for DENV-2: five sera negative for DENV-2 and yellow fever virus (YFV), five sera negative for antibodies against DENV-2 and positive for antibodies against YFV, and five sera positive for antibodies against DENV-2 that were selected using the plate reduction neutralization test. Results: The optimum method for MNT used suspended cells (0.9 X 105 cells/mL), 2% fetal bovine serum, and viral particles at the 5th passage. The minimal dilution able to differentiate a positive sample for DENV2 was 1:40, and the MNT incubation time for DENV-2 was ten days. Conclusion: MNT with the cell suspension method and a culture medium with 2% fetal bovine serum allows the detection of IgG neutralizing antibodies against DENV-2 with reliable results, so that larger sample sizes may be assessed, saving materials to be used.
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Abstract Snakebite envenoming is a significant global health challenge, and for over a century, traditional plasma-derived antivenoms from hyperimmunized animals have been the primary treatment against this infliction. However, these antivenoms have several inherent limitations, including the risk of causing adverse reactions when administered to patients, batch-to-batch variation, and high production costs. To address these issues and improve treatment outcomes, the development of new types of antivenoms is crucial. During this development, key aspects such as improved clinical efficacy, enhanced safety profiles, and greater affordability should be in focus. To achieve these goals, modern biotechnological methods can be applied to the discovery and development of therapeutic agents that can neutralize medically important toxins from multiple snake species. This review highlights some of these agents, including monoclonal antibodies, nanobodies, and selected small molecules, that can achieve broad toxin neutralization, have favorable safety profiles, and can be produced on a large scale with standardized manufacturing processes. Considering the inherent strengths and limitations related to the pharmacokinetics of these different agents, a combination of them might be beneficial in the development of new types of antivenom products with improved therapeutic properties. While the implementation of new therapies requires time, it is foreseeable that the application of biotechnological advancements represents a promising trajectory toward the development of improved therapies for snakebite envenoming. As research and development continue to advance, these new products could emerge as the mainstay treatment in the future.
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Snake Bites/drug therapy , Antivenins/therapeutic use , SnakesABSTRACT
@#ObjectiveTo develop and apply a method for detecting the titer of varicella-zoster virus(VZV)neutralizing antibodies based on complement dependence,so as to improve the sensitivity of traditional plaque reduction neutralization assay for detection of the titer of VZV antibody.MethodsThe antigen(live attenuated varicella vaccine)and antibody(human VZV immunoglobulin)were mixed in different proportions and different incubation times. After neutralization,the antigen-antibody mixture was inoculated into human diploid cell 2BS strain cultured in a six-well plate. After 7 ～ 10 d of culture,the number of plaques was counted by Coomassie brilliant blue staining,and the 50% neutralizing antibody titer was calculated by Karber′s formula. Under the optimal neutralization conditions obtained,the effect of complement on the sensitivity of neutralization experiment was explored by changing the addition amount of complement(lyophilized guinea pig serum)to evaluate the optimal addition amount of complement. According to the determined neutralization test parameters,the neutralizing antibody titers of 12 anti-VZV mouse sera and 14 anti-VZV human sera were detected by using traditional plaque method and complement-dependent plaque method respectively.ResultsThe key parameters of the detection method were determined:the titer of VZV standard antigen was 500 ～ 1 000 PFU/mL;the proportion of complement added to the antigen-antibody neutralization system was 1∶10(v/v),and the neutralization condition was 37 ℃ for 1 h. Both the complement-dependent plaque method and the traditional plaque method were positive for anti-VZV mouse serum antibody,while the antibody titer detected by the traditional plaque method was generally lower,and the antibody level of mice inoculated with 2 doses of live attenuated varicella vaccine was significantly higher than that of mice inoculated with 1 dose(t = 0. 45,P < 0. 05);Both of the two methods were positive for anti-VZV human serum antibody.ConclusionA complement-dependent detection method for neutralizing antibody titer of VZV was established. The addition of complement significantly improved the sensitivity of neutralization detection. The evaluation of the titers of neutralizing antibodies in mouse serum with different immunization strategies by the method suggested that the immune effect of two doses of vaccine was better than that of one dose.
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@#Objective To compare three methods for detection of antibody level in serum immunized with SARS-CoV-2mRNA vaccine. Methods Enzyme-linked immunosorbent assay(ELISA),pseudo virus-based neutralization assay(PBNA)and micro-cytopathic effect neutralization test(MCPENT)were used to detect the antibody levels of a total of 120 serum samples(40 before immunization and 80 after immunization)before and after 2 doses of mRNA vaccine immunization,and the consistency and correlation of the three methods were analyzed. Results The consistency rates of the three methods detecting 120 serum samples were all over 90%,the Kappa coefficients were all more than 0. 7,and each P was less than0. 01. The correlation coefficient(r)between the antibody potency results of positive serum samples detected by the three methods was 0. 825～0. 902,and each P was less than 0. 01. Conclusion The three methods have good consistency and correlation in detecting antibody level of serum immunized with SARS-CoV-2 mRNA vaccine.
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@#Objective To prepare the second generation internal control reference(B2)for Ig G antibody against severe acute respiratory symptom coronavirus 2(SARS-CoV-2)and evaluate its applicability in ELISA detection method. Methods Among the volunteers vaccinated with SARS-CoV-2 inactivated vaccine(BBIBP-Cor V)produced by Beijing Institute of Biological Products Co.,Ltd.,19 Ig G antibody positive plasma samples with ELISA-Ig G dilution ratio of 20 ～ 60 were screened,and the Ig G antibody,IgM antibody and neutralizing antibody were detected by ELISA,B2 was prepared from nonlipid plasma with ELISA-Ig G dilution ratio of 32 ～ 45,IgM negative and similar neutralizing antibody inhibition rate. The neutralizing antibody potency of the first generation internal control reference(B1)and B2 detected by ELISA was calibrated with the first generation WHO international standard of anti-SARS-CoV-2 immunoglobulin(NIBSC 20/136),and the accelerated stability(storage at 2 ～ 8 ℃ for 5,8,14,20,and 30 d respectively),the service stability(storage at 18 ～25 ℃ for 1,2,and 3 h respectively),the freeze-thaw stability(1,2 and 3 times)and the long-term stability(storage at-25 ℃ for10 months)of B2 were tested. B2 was used as standard to detect plasma after single vaccine immunization and mixed plasma was prepared according to different ELISA-Ig G dilution ratio. The correlation and linear regression analysis between ELISA-Ig G dilution ratio and neutralizing antibody potency of pseudovirus in mixed plasma were carried out. Results Among 19 plasma samples,5 samples were non-lipid plasma with ELISA-Ig G dilution ratio of 32 ～ 45,IgM negative and similar neutralizing antibody inhibition rate. B2 was prepared by mixing every plasma in equal volume fraction,and the dilution ratio of ELISA-Ig G was assigned to 32. The neutralizing antibody potency of B1 calibrated with NIBSC 20/136 was 133. 38 EIU/m L and that of B2 was 122. 14 EIU/m L. The recovery rates of accelerated stability,service stability,freeze-thaw stability and long-term stability of B2 were all in the range of(100 ± 15)%. The ELISA-Ig G dilution ratio of the mixed plasma from the same source was significantly correlated with the neutralizing antibody potency of pseudovirus.(each R~2> 0. 99,each P < 0. 000 1).Conclusion B2 prepared from plasma immunized with SARS-CoV-2 inactivated vaccine can replace B1 prepared from plasma of COVID-19 convalescent patients.
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Objective:To establish an in vivo infection model of H5N1 pseudovirus and to detect the neutralizing activity of FHA3 antibody using this model. Methods:Based on the sequence information of hemagglutinin (HA) and neuraminidase (NA) of A/Anhui/1/2005/H5N1 strain, two recombinant plasmids of pcDNA3.1-HA5 and pcDNA3.1-NA1 were constructed. The two plasmids and plasmid pNL4-3.Luc.R-E- were co-transfected into 293T cells to prepare H5N1 pseudovirus supernatant. The morphology of pseudovirus particles in the supernatant was observed by electron microscopy. MDCK cells were infected with the pseudovirus supernatant and the virus titer was detected. BALB/c mice were injected with the pseudovirus supernatant by intraperitoneal injection and subjected to bioluminescence imaging at 2, 5, 8, and 12 d after infection to detect the pseudovirus infection in vivo. The functional activity of FHA3 antibody in vivo was evaluated using the established mouse infection model. Results:The recombinant plasmids pcDNA3.1-HA5 and pcDNA3.1-NA1 were correctly constructed and could be used to prepare pseudovirus supernatants of high titer by co-transfecting 293T cells with the plasmid pNL4-3.Luc.R-E-. The virus particles were round under electron microscope. H5N1 pseudovirus-infected mice exhibits strong fluorescence signals, which were attenuated by FHA3 treatment before challenge.Conclusions:The in vivo infection model of H5N1 pseudovirus was successfully constructed and FHA3 antibody was proved to be protective against the pseudovirus infection.
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There are currently eight vaccines against SARS-CoV-2 that have received Emergency Use Authorization by the WHO that can offer some protection to the world’s population during the COVID-19 pandemic. Though research is being published all over the world, public health officials, policymakers and governments are collecting evidence-based information to establish the public health policies. Unfortunately, continued international travel, violations of lockdowns and social distancing, the lack of mask use, the emergence of mutant strains of the virus and lower adherence by a sector of the global population that remains sceptical of the protection offered by vaccines, or about any risks associated with vaccines, hamper these efforts. Here we examine the literature on the efficacy, effectiveness and safety of COVID-19 vaccines, with an emphasis on select categories of individuals and against new SARS-CoV-2 strains. The literature shows that these eight vaccines are highly effective in protecting the population from severe disease and death, but there are some issues concerning safety and adverse effects. Further, booster shots and variant-specific vaccines would also be required.
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The WHO emergency use-listed (EUL) COVID-19 vaccines were developed against early strains of SARS-CoV-2. With the emergence of SARS-CoV-2 variants of concern (VOCs) - Alpha, Beta, Gamma, Delta and Omicron, it is necessary to assess the neutralizing activity of these vaccines against the VOCs. PubMed and preprint platforms were searched for literature on neutralizing activity of serum from WHO EUL vaccine recipients, against the VOCs, using appropriate search terms till November 30, 2021. Our search yielded 91 studies meeting the inclusion criteria. The analysis revealed a drop of 0-8.9-fold against Alpha variant, 0.3-42.4-fold against Beta variant, 0-13.8-fold against Gamma variant and 1.35-20-fold against Delta variant in neutralization titres of serum from the WHO EUL COVID-19 vaccine recipients, as compared to early SARS-CoV-2 isolates. The wide range of variability was due to differences in the choice of virus strains selected for neutralization assays (pseudovirus or live virus), timing of serum sample collection after the final dose of vaccine (day 0 to 8 months) and sample size (ranging from 5 to 470 vaccinees). The reasons for this variation have been discussed and the possible way forward to have uniformity across neutralization assays in different laboratories have been described, which will generate reliable data. Though in vitro neutralization studies are a valuable tool to estimate the performance of vaccines against the backdrop of emerging variants, the results must be interpreted with caution and corroborated with field-effectiveness studies.
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Abstract Snake envenomation is a public health problem, and while serum therapy prevents death, the local effects of venoms can lead to amputations or morbidities. Thus, alternative treatments deserve attention. In this study, we tested eight derivatives of 1,2,3-triazole against some toxic activities of Bothrops jararaca venom. The derivatives were synthesized, and their structures analyzed by infrared and nuclear magnetic resonance. After that, the ability of compounds to inhibit hemolysis, coagulation, proteolysis, hemorrhaging, edema, and lethal activities of B. jararaca venom was investigated. The derivatives were incubated with B. jararaca venom (incubation protocol), administered before (prevention protocol) or after (treatment protocol) injecting venom into the mice. Then, hemorrhaging assay occurred. As a result, most of the derivatives inhibited the activities, even if they were incubated, injected before or after B. jararaca venom. However, the derivatives TRI 07 and TRI 18 seemed to be the most efficient in impairing hemorrhaging. The derivatives showed a low drug score of toxicity based on an in silico technique. Therefore, the derivatives fulfilled physicochemical and biological requirements to become drugs, and they may be a brand new initiative for designing antivenom molecules to complement antivenom therapy to efficiently block tissue necrosis or any other local effects.
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Objective:To study the protective effects of bicistronic DNA vaccines carrying herpes simplex virus type 2 glycoprotein D (HSV-2 gD) and adjuvant CCL28 sequences that were connected by internal ribosome entry site (IRES) sequence in mouse model.Methods:The recombinant DNA vaccines, pgD-IRES-CCL28 and pCCL28-IRES-gD, encoding HSV-2 gD and adjuvant CCL28 were constructed with IRES sequence. After verified by sequencing, they were intramuscularly injected twice into BALB/c mice. Serum samples and vaginal lavage fluids were collected regularly. Splenocytes, mesenteric lymph node cells and rectal mucosa tissues were separated and collected. The titers of antigen-specific antibodies in immunized mice were analyzed with end-point ELISA. In vitro neutralization assay was used to measure neutralizing antibody titers in serum and vaginal lavage fluid after vaccination and virus challenge. CCL28-responsive immune cells in splenocytes, mesenteric lymph node cells and rectal tissues were detected by chemotaxis experiment and immunohistochemical staining. The protective effects of the bicistronic DNA vaccines were evaluated by fluorescent quantitative PCR, weighing and disease severity assessment. Humoral and cellular immune responses induced by the bicistronic DNA vaccines and their efficacy in immunoprotection were analyzed by comparing with pgD+ pCCL28 group. Results:IgG titers in serum samples and IgA antibody titers in vaginal lavage fluids of mice immunized with pCCL28-IRES-gD were similar to those in pgD+ pCCL28 group. The neutralizing ability of antibodies, the number of rectal mucosal IgA+ plasma cells and CCL28-responsive immune cells in mucosal tissues were increased in pCCL28-IRES-gD group. Serum neutralizing antibodies were not produced immediately in the mice challenged with HSV-2, but no weight loss, disease symptoms or death was observed. However, pgD+ pcDNA3.1 and pgD-IRES-CCL28 were ineffective against HSV-2 infection in mice.Conclusions:The recombinant bicistronic DNA vaccine of pCCL28-IRES-gD could induce stronger mucosal immune response in mice and provide better protective effects.
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Objective:To compare the immune responses to simply mixed and fused recombinant DNA vaccines of herpes simplex virus type 2 glycoprotein D (HSV-2 gD) and molecular adjuvant CCL19 in mice and to evaluate the protective effects.Methods:Gene recombination technology was used to construct recombinant DNA vaccines expressing HSV-2 gD and CCL19 alone or fused together. After verification by sequencing, Western blot and ELISA, BALB/c mice were immunized twice by intramuscular injection. Serum samples and vaginal lavage fluids were collected regularly after immunization. Splenocytes, mesenteric lymph node cells and rectal tissues were collected after immunization. Differences in humoral and cellular immune responses to the two forms of vaccines and their protective effects in mice were analyzed using end-point ELISA, in vitro neutralization assay, immunohistochemical staining, chemotaxis assay, vaginal virus challenge, fluorescence quantitative PCR, weighing and disease severity assessment. Results:The fused recombinant pgD-IZ-CCL19 plasmid could express gD protein and CCL19 protein in vitro, but the level of expressed CCL19 protein by pCCL19-IZ-gD plasmid was less than that by pgD-IZ-CCL19. The mice immunized with pgD-IZ-CCL19 showed higher levels of IgG in sera and IgA in vaginal lavage fluids ( P<0.01) and stronger neutralization ability than the mice vaccinated with pgD+ pCCL19. Compared with other groups, more lymphocytes were recruited in the rectal mucosa, the spleen and mesenteric lymph nodes of mice immunized with pgD-IZ-CCL19. Weight loss or disease symptoms were not observed in the pgD-IZ-CCL19 group after virus challenge. In addition, the positive rate of HSV-2 in vaginal mucosa and the mortality rate in the pgD-IZ-CCL19 group were the lowest. However, pCCL19-IZ-gD turned out ineffective in preventing HSV-2 infection. Conclusions:The fused recombinant DNA vaccine pgD-IZ-CCL19 could induce stronger immune responses in mice and provide better protective effects, which was superior to the simply mixed DNA vaccine.
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【Objective】 To assess three severe acute respiratorysyndrome coronavirus 2 (SARS-CoV-2) enzyme linked immunosorbent assays (ELISA) and one pseudotype lentivirus-based neutralization test (ppNAT) in detecting the convalescent plasma antibody levles from COVID-19. 【Methods】 30 COVID-19 convalescent plasma samples were screened for antibodies against SARS-CoV-2 using three kinds of SARS-CoV-2 ELISA reagents and one ppNAT test in Shenzhen. The controls consisted of plasma samples from 32 healthy blood donors in February 2019. The diagnostic efficacy analysis of various SARS-CoV-2 ELISA reagents was performed using real-time fluorescent Polymerase Chain Reaction (RT-PCR). We also analyzed correlation between different immunological reagents and the age, gender, hospitalization, and severity of illness. 【Results】 The positive yielding rate of ppNAT and three kinds of IgG ELISA was higher than that of IgM ELISA. The positive yielding rates of three kinds of IgG ELISA were 100%(30/30), 93.33%(28/30), and 96.67%(29/30) respectively, while the yielding rates in control group were all 0. The positive yielding rate of three IgM ELISAs were 93.33%(28/30), 70%(21/30)and 46.67% (14/30). All the cases from negative control group were negative for IgG and IgM. Pearson correlation coefficient was calculated; there was a strong correlation between ELISA reagent 2 IgG and ELISA reagent 3 IgG (r=0.765, P0.05). 【Conclusion】 In the convalescent plasma with nucleic acid confirmed covid-19, the yielding rates of different IgM antibodies varied greatly. Antibody levels were influenced by age to some extent.
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Objective:To improve the consistency of test results through reducing inter-laboratory variation in SARS-CoV-2 antibody detection with WHO SARS-CoV-2 antibody candidate international standard (IS, sample G) and antibody reference panel (samples E, F, H, I, J).Methods:Ten WHO samples (A-J) including the candidate IS and reference panel were evaluated using different methods, such as microneutralization tests based on live SARS-CoV-2, pseudovirus neutralization assay and commercial ELISA kits. The test results were compared using statistical analysis.Results:Using IS (sample G) as a reference, the relative concentrations of other samples could be determined with less variation. ELISA and pseudovirus neutralization assay had consistent results with those obtained with the microneutralization test based on SARS-COV-2 strain HB02. Weakly positive samples could be detected only by a certain kit.Conclusions:The availability of an IS for antibodies would facilitate the standardization of SARS-CoV-2 antibody detection methods. The reference panel fitted all the assays based on the SARS-CoV-2 prototype Wuhan strain. Pseudovirus neutralization assay and ELISA could be used as alternatives to live SARS-CoV-2-based neutralization test to some extent.
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Objective:To evaluate the performance of two commercial EIA kits for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) neutralizing antibodies.Methods:Two commercial SARS-CoV-2 neutralizing antibody ELISA test kits (A and B) were used to detect serum panel consists of the following sera: 44 collected before vaccination, 120 collected one month after vaccination and 64 collected six months after recovery from convalescent patients of COVID-19. In the meantime, the above samples were also taken for live virus micro-neutralization test (micro-NT) indicated as the 50% neutralization antibody titer (NT 50). The consistency of qualitative and quantitative results between the two commercial kits and live virus neutralization test was analyzed. Results:Taking the micro-NT results as the standard, the positive coincidence rates of A and B kits were 97.40% and 100.00%, respectively; the negative coincidence rates were 97.30% and 95.95%, respectively; the Youden indices were 0.95 and 0.96, respectively. Furthermore, quantitative analysis indicated that the correlation coefficients between A and B kits and micro-NT results were 0.24 ( P<0.05) and 0.52 ( P<0.000 1) for samples collected after vaccination, respectively; while the correlation coefficients were 0.81 ( P<0.000 1) and 0.89 ( P<0.000 1) for convalescent serum samples, respectively. Conclusions:The results obtained by the two commercial neutralizing antibody detection kits were in good agreement with the qualitative results of micro-NT. The neutralizing antibody titers in convalescent serum samples detected by the two kits showed a stronger correlation with the micro-NT results.
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Objective:To evaluate the in vitro cross-neutralization of serum antibodies in human and mice immunized with inactivated SARS-CoV-2 vaccine against Delta and Beta variants. Methods:Human serum samples after a second and a third dose of inactivated SARS-CoV-2 vaccine and mouse serum samples after a two-dose vaccination were collected. The neutralizing antibodies in the samples against SARS-CoV-2 strains of prototype, Delta and Beta variants were detected using micro-neutralization assay in biosafety level Ⅲ laboratory. The seroconversion rates and geometric mean titers (GMTs) of antibodies were calculated.Results:The seroconversion rates of antibodies in human serum samples against different SARS-CoV-2 strains were all above 95%. After two-dose vaccination, the GMTs of neutralizing antibodies against the prototype, Delta and Beta strains were 109, 41 and 15, respectively. The GMTs decreased by 2.7 folds and 7.3 folds for the Delta and Beta variants as compared with the prototype strain. After the booster vaccination, the GMTs of neutralizing antibodies against the prototype, Delta and Beta strains were 446, 190 and 86, respectively. The GMTs of neutralizing antibodies against Delta and Beta variants decreased by 2.3 folds and 5.2 folds as compared with that against the prototype strain. The seroconversion rates of antibodies against different SARS-CoV-2 strains in mouse serum samples were all 100%. The GMTs of neutralizing antibodies against the prototype, Delta and Beta strains were 2 037, 862 and 408, respectively. The GMTs decreased by 2.4 folds and 5.0 folds for the Delta and Beta variants.Conclusions:Inactivated SARS-CoV-2 vaccine could induce a certain level of neutralizing antibodies against Delta and Beta variants in both human and mouse models. Moreover, a third dose of vaccine induced higher levels of neutralizing antibodies against Delta and Beta variants in human. This study provided valuable data for the clinical application and protective evaluation of the inactivated SARS-CoV-2 vaccine.
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Objetivou-se investigar a presença do Vírus da Estomatite Vesicular (VEV) e seus fatores de risco para ocorrência e disseminação da enfermidade em equídeos das mesorregiões Leste e Oeste Potiguar do estado do Rio Grande do Norte, Brasil. Foram analisadas pela técnica de virusneutralização, 809 amostras sanguíneas de equídeos provenientes de noventa propriedades de dezesseis municípios Potiguares durante os meses de julho de 2018 a fevereiro de 2019. Os fatores de riscos associados ao VEV foram avaliados por meio de questionário epidemiológico e os dados submetidos a análise estatística no programa IBM SPSS Statistics versão 21.0 com nível de confiança de 95%. Posteriormente, todas as variáveis estatisticamente significantes foram submetidas a análise de regressão de Poisson. A soroprevalência de anticorpos anti-VEV foi 24,6% (199/809), sendo 3,2% (13/402) de soropositivos na mesorregião Leste e 45,7% (186/407) na do Oeste Potiguar. Com relação aos sorotipos, observou-se uma prevalência de 3,8% (31/809) e 24,5% (198/809) para Indiana 2 e 3 respectivamente, com 15,1% (30/198) de coinfecção. Equídeos criados na mesorregião Oeste, em propriedades que não realizam quarentena e onde os animais enfermos são mantidos no rebanho, foram consideradas fatores predisponentes a infecção pelo VEV. Esses resultados demonstram a circulação do VEV em equídeos no Rio Grande do Norte, com destaque ao Oeste Potiguar, e sendo necessário a aplicação de medidas sanitárias que impeçam introdução e disseminação do vírus ente as espécies susceptíveis, principalmente em condições climáticas favoráveis para a sua manutenção, no ambiente de criação e pastagens.
This study aimed to investigate the presence of Vesicular stomatitis virus (VSV) and risk factors for its occurrence and dissemination in equines from the Eastern and Western mesoregions of the state of Rio Grande do Norte, Brazil. Blood samples were analyzed, by Serum Virus Neutralization Assay, from 809 animals belonging to 90 properties distributed in sixteen municipalities from July 2018 to February 2019. Risk factors were assessed using an epidemiological questionnaire. Data were submitted to statistical analysis using the software IBM SPSS Statistics, version 21.0 with a 95% confidence level. Also, all statistically significant variables were subjected to Poisson regression analysis. The occurrence of anti-VSV antibodies was 24.6% (199/809) with 3.2% (13/402) and 45.7% (186/407) of seropositivity in the Western and Eastern mesoregion, respectively. Regarding serotypes, there were an occurrence of 3.8% (31/809) and 24.5% (198/809) for Indiana 2 and 3, respectively, and 15.1% (30/198) of co-infection for both. Equines kept of the Western mesoregion, on properties that do not quarantine, and where sick animals are kept in the herd, were considered risk factors for LVV infection. These results demonstrate the presence of VSV in equines in Rio Grande do Norte, with emphasis on Oeste Potiguar, and that sanitary measures must be adopted to prevent the introduction and viral spreading among susceptible species, especially due to favorable climatic conditions for the maintenance of VSV in the breeding and pasture environment.
Subject(s)
Animals , Horses , Horses/virology , Vesicular Stomatitis/virology , Biological Factors/analysis , Risk Factors , Rhabdoviridae Infections/diagnosisABSTRACT
Objetivou-se investigar a presença do Vírus da Estomatite Vesicular (VEV) e seus fatores de risco para ocorrência e disseminação da enfermidade em equídeos das mesorregiões Leste e Oeste Potiguar do estado do Rio Grande do Norte, Brasil. Foram analisadas pela técnica de virusneutralização, 809 amostras sanguíneas de equídeos provenientes de noventa propriedades de dezesseis municípios Potiguares durante os meses de julho de 2018 a fevereiro de 2019. Os fatores de riscos associados ao VEV foram avaliados por meio de questionário epidemiológico e os dados submetidos a análise estatística no programa IBM SPSS Statistics versão 21.0 com nível de confiança de 95%. Posteriormente, todas as variáveis estatisticamente significantes foram submetidas a análise de regressão de Poisson. A soroprevalência de anticorpos anti-VEV foi 24,6% (199/809), sendo 3,2% (13/402) de soropositivos na mesorregião Leste e 45,7% (186/407) na do Oeste Potiguar. Com relação aos sorotipos, observou-se uma prevalência de 3,8% (31/809) e 24,5% (198/809) para Indiana 2 e 3 respectivamente, com 15,1% (30/198) de coinfecção. Equídeos criados na mesorregião Oeste, em propriedades que não realizam quarentena e onde os animais enfermos são mantidos no rebanho, foram consideradas fatores predisponentes a infecção pelo VEV. Esses resultados demonstram a circulação do VEV em equídeos no Rio Grande do Norte, com destaque ao Oeste Potiguar, e sendo necessário a aplicação de medidas sanitárias que impeçam introdução e disseminação do vírus ente as espécies susceptíveis, principalmente em condições climáticas favoráveis para a sua manutenção, no ambiente de criação e pastagens.
This study aimed to investigate the presence of Vesicular stomatitis virus (VSV) and risk factors for its occurrence and dissemination in equines from the Eastern and Western mesoregions of the state of Rio Grande do Norte, Brazil. Blood samples were analyzed, by Serum Virus Neutralization Assay, from 809 animals belonging to 90 properties distributed in sixteen municipalities from July 2018 to February 2019. Risk factors were assessed using an epidemiological questionnaire. Data were submitted to statistical analysis using the software IBM SPSS Statistics, version 21.0 with a 95% confidence level. Also, all statistically significant variables were subjected to Poisson regression analysis. The occurrence of anti-VSV antibodies was 24.6% (199/809) with 3.2% (13/402) and 45.7% (186/407) of seropositivity in the Western and Eastern mesoregion, respectively. Regarding serotypes, there were an occurrence of 3.8% (31/809) and 24.5% (198/809) for Indiana 2 and 3, respectively, and 15.1% (30/198) of co-infection for both. Equines kept of the Western mesoregion, on properties that do not quarantine, and where sick animals are kept in the herd, were considered risk factors for LVV infection. These results demonstrate the presence of VSV in equines in Rio Grande do Norte, with emphasis on Oeste Potiguar, and that sanitary measures must be adopted to prevent the introduction and viral spreading among susceptible species, especially due to favorable climatic conditions for the maintenance of VSV in the breeding and pasture environment.
Subject(s)
Animals , Vesicular stomatitis Indiana virus , Horse Diseases/virology , Risk Factors , Vesicular Stomatitis/virology , Antibodies, Viral/analysisABSTRACT
Biodiesel is a clean and cyclical energy resource that is derived from animal and/or vegetable fat. As it blends well with petrodiesel, biodiesel is added to Brazilian commercial diesel. The main raw materials used to produce biodiesel in Brazil include soybean, corn, and sunflower oils. However, these are also used for human consumption and hence, have a high market value. Therefore, pinhão manso oil, which exhibits high productivity at low cost, is a promising alternative. However, the high acidity index of this oil results in a low transesterification yield and the produced biofuel does not meet the requirements imposed by the ANP. Thus, this study intends to demonstrate that a large part of the free fatty acids in pinion oil are present in the seed endocarp. For the development of the project, the oil was extracted by hot solvent, using the soxhlet equipment and the hexane solvent, to determine the acidity index, the titration technique was used, the titrant used was sodium hydroxide. So the acidity index of the oil extracted from the seed with its shell is 10.9 mgKOH/g, while the lipid obtained without the shell exhibits a value of 0.95 mgKOH/g, proving the influence of the endocarp.
Subject(s)
Seeds , Plant Oils , Jatropha , Fatty AcidsABSTRACT
The present research evaluated the seroprevalence of anti-zika virus (anti-ZIKV) antibodies by virus neutralization test (VNT) in 529 bovines from Andradina city, São Paulo state, Brazil. The reading was performed in an inverted optical microscope, considering reagents when the antibodies were capable to neutralize the ZIKV. Of the 529 samples, 53 (10.01%) were reagents. The animals were healthy at the time of collection. The samples were collected in February 2018, a favorable period for the multiplication of the vector and the highest risk of disease transmission. None of the animals showed anti-bovine viral diarrhea virus (anti-BVDV) antibodies, ruling out a possible cross-reaction, reinforcing the possible contact of the bovine with the ZIKV. In the herd, 88 pregnant females were evaluated; of these, 12 cows were reactive, with no history of reproductive problems or fetal malformations. This is the first research on the seroprevalence of ZIKV in cattle in Brazil, and studies should continue to evaluate cattle as a possible host of this arbovirus and its possible consequences for unique health and agribusiness.