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BACKGROUND: Middle East Respiratory Syndrome Coronavirus is a highly pathogenic virus that poses a significant threat to public health. OBJECTIVE: The purpose of this study is to develop and characterize novel mouse monoclonal antibodies targeting the spike protein S1 subunit of the Middle East Respiratory Syndrome Corona Virus (MERS-CoV). METHODS: In this study, three mouse monoclonal antibodies (mAbs) against MERS-CoV were generated and characterized using hybridoma technology. The mAbs were evaluated for their reactivity and neutralization activity. The mAbs were generated through hybridoma technology by the fusion of myeloma cells and spleen cells from MERS-CoV-S1 immunized mice. The resulting hybridomas were screened for antibody production using enzyme-linked immunosorbent assays (ELISA). RESULTS: ELISA results demonstrated that all three mAbs exhibited strong reactivity against the MERS-CoV S1-antigen. Similarly, dot-ELISA revealed their ability to specifically recognize viral components, indicating their potential for diagnostic applications. Under non-denaturing conditions, Western blot showed the mAbs to have robust reactivity against a specific band at 116 KDa, corresponding to a putative MERS-CoV S1-antigen. However, no reactive bands were observed under denaturing conditions, suggesting that the antibodies recognize conformational epitopes. The neutralization assay showed no in vitro reactivity against MERS-CoV. CONCLUSION: This study successfully generated three mouse monoclonal antibodies against MERS-CoV using hybridoma technology. The antibodies exhibited strong reactivity against MERS-CoV antigens using ELISA and dot ELISA assays. Taken together, these findings highlight the significance of these mAbs for potential use as valuable tools for MERS-CoV research and diagnosis (community and field-based surveillance and viral antigen detection).
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Anticorpos Monoclonais , Anticorpos Neutralizantes , Anticorpos Antivirais , Ensaio de Imunoadsorção Enzimática , Hibridomas , Coronavírus da Síndrome Respiratória do Oriente Médio , Glicoproteína da Espícula de Coronavírus , Animais , Glicoproteína da Espícula de Coronavírus/imunologia , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Anticorpos Monoclonais/imunologia , Camundongos , Anticorpos Antivirais/imunologia , Hibridomas/imunologia , Humanos , Anticorpos Neutralizantes/imunologia , Camundongos Endogâmicos BALB C , Epitopos/imunologia , Testes de Neutralização , Especificidade de Anticorpos/imunologia , Infecções por Coronavirus/imunologia , FemininoRESUMO
Antibiotic resistant bacteria are a growing threat to global health security. Whilst the emergence of antimicrobial resistance (AMR) is a natural phenomenon, it is also driven by antibiotic exposure in health care, agriculture, and the environment. Antibiotic pressure and inappropriate use of antibiotics are important factors which drive resistance. Apart from their use to treat bacterial infections in humans, antibiotics also play an important role in animal husbandry. With limited antibiotic options, alternate strategies are required to overcome AMR. Passive immunization through oral, nasal and topical administration of egg yolk-derived IgY antibodies from immunized chickens were recently shown to be effective for treating bacterial infections in animals and humans. Immunization of chickens with specific antigens offers the possibility of creating specific antibodies targeting a wide range of antibiotic-resistant bacteria. In this review, we describe the growing global problem of antimicrobial resistance and highlight the promising potential of the use of egg yolk IgY antibodies for the treatment of bacterial infections, particularly those listed in the World Health Organization priority list.
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Anti-Infecciosos , Infecções Bacterianas , Humanos , Animais , Galinhas , Anticorpos , Infecções Bacterianas/prevenção & controle , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Resistência Microbiana a MedicamentosRESUMO
Safe, passive immunization methods are required against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and its variants. Immunization of chickens with antigen is known to induce specific IgY antibodies concentrated in the egg yolk and has a good safety profile, high yield of IgY per egg, can be topically applied, not requiring parenteral delivery. Our data provide the first evidence of the prophylactic efficacy of Immunoglobulin Y antibodies against SARS-CoV-2 in mice. Lohmann hens were injected with recombinant SARS-CoV-2 RBD protein; IgY-Abs were extracted from the eggs and characterized using SDS-PAGE. Antiviral activity was evaluated using plaque reduction neutralization tests. In additional experiments, IgY-RBD efficacy was examined in mice sensitized to SARS-CoV-2 infection by transduction with Ad5-hACE2 (mild disease) or by using mouse-adapted virus (severe disease). In both cases, prophylactic intranasal administration of IgY-Abs reduced SARS-CoV-2 replication, and reduced morbidity, inflammatory cell infiltration, hemorrhage, and edema in the lungs and increased survival compared to control groups that received non-specific IgY-Abs. These results indicate that further evaluation of IgY-RBD antibodies in humans is warranted.
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COVID-19 , SARS-CoV-2 , Animais , Anticorpos Antivirais , Antivirais , COVID-19/prevenção & controle , Galinhas , Feminino , Humanos , Imunoglobulinas , CamundongosRESUMO
Identified in 2012, the Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe and often fatal acute respiratory illness in humans. No approved prophylactic or therapeutic interventions are currently available. In this study, we developed chicken egg yolk antibodies (IgY Abs) specific to the MERS-CoV spike (S) protein and evaluated their neutralizing efficiency against MERS-CoV infection. S-specific IgY Abs were produced by injecting chickens with the purified recombinant S protein of MERS-CoV at a high titer (4.4 mg/mL per egg yolk) at week 7 post immunization. Western blotting and immune-dot blot assays demonstrated specific binding to the MERS-CoV S protein. In vitro neutralization of the generated IgY Abs against MERS-CoV was evaluated and showed a 50% neutralizing concentration of 51.42 µg/mL. In vivo testing using a human-transgenic mouse model showed a reduction of viral antigen positive cells in treated mice, compared to the adjuvant-only controls. Moreover, the lung cells of the treated mice showed significantly reduced inflammation, compared to the controls. Our results show efficient neutralization of MERS-CoV infection both in vitro and in vivo using S-specific IgY Abs. Clinical trials are needed to evaluate the efficiency of the IgY Abs in camels and humans.
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BACKGROUND: The unusual pneumonia outbreak that originated in the city of Wuhan, China in December 2019 was found to be caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), or COVID-19. METHODS: In this work, we have performed an in silico design and prediction of potential siRNAs based on genetic diversity and recombination patterns, targeting various genes of SARS-CoV-2 for antiviral therapeutics. We performed extensive sequence analysis to analyze the genetic diversity and phylogenetic relationships, and to identify the possible source of virus reservoirs and recombination patterns, and the evolution of the virus as well as we designed the siRNAs which can be used as antivirals against SARS-CoV-2. RESULTS: The sequence analysis and phylogenetic relationships indicated high sequence identity and closed clusters with many types of coronavirus. In our analysis, the full-genome of SARS-CoV-2 showed the highest sequence (nucleotide) identity with SARS-bat-ZC45 (87.7%). The overall sequence identity ranged from 74.3% to 87.7% with selected SARS viruses. The recombination analysis indicated the bat SARS virus is a potential recombinant and serves as a major and minor parent. We have predicted 442 siRNAs and finally selected only 19 functional, and potential siRNAs. CONCLUSION: The siRNAs were predicted and selected based on their greater potency and specificity. The predicted siRNAs need to be validated experimentally for their effective binding and antiviral activity.
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COVID-19 , SARS-CoV-2 , Antivirais/farmacologia , Simulação por Computador , Humanos , Filogenia , RNA Interferente Pequeno/genéticaRESUMO
The Middle East respiratory syndrome coronavirus (MERS-CoV) was identified in 2012 and causes severe and often fatal acute respiratory illness in humans. No approved prophylactic and therapeutic interventions are currently available. In this study, we have developed egg yolk antibodies (immunoglobulin Y (IgY)) specific for MERS-CoV spike protein (S1) in order to evaluate their neutralizing efficiency against MERS-CoV infection. S1-specific immunoglobulins were produced by injecting chickens with purified recombinant S1 protein of MERS-CoV at a high titer (5.7 mg/mL egg yolk) at week 7 post immunization. Western blotting and immune-dot blot assays demonstrated that the IgY antibody specifically bound to the MERS-CoV S1 protein. Anti-S1 antibodies were also able to recognize MERS-COV inside cells, as demonstrated by an immunofluorescence assay. Plaque reduction and microneutralization assays showed the neutralization of MERS-COV in Vero cells by anti-S1 IgY antibodies and non-significantly reduced virus titers in the lungs of MERS-CoV-infected mice during early infection, with a nonsignificant decrease in weight loss. However, a statistically significant (p = 0.0196) quantitative reduction in viral antigen expression and marked reduction in inflammation were observed in lung tissue. Collectively, our data suggest that the anti-MERS-CoV S1 IgY could serve as a potential candidate for the passive treatment of MERS-CoV infection.
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The protective effects of both manuka and talh honeys were assessed using a rat model of cisplatin (CISP)-induced hepatotoxicity and nephrotoxicity. The results revealed that both honeys exerted a protective effect against CISP-induced hepatotoxicity and nephrotoxicity as demonstrated by decreasing liver and kidney function. Manuka honey also prevented CISP-induced histopathological changes observed in the liver and decreased the changes seen in the kidneys. Talh honey decreased CISP-induced liver histopathological changes but had no effect on CISP-induced kidney histopathological changes. Both honeys reduced the oxidative stress in the liver. Conversely, they have no effect on kidney oxidative stress, except that manuka honey increased CAT activity. GC-MS analysis showed the presence of the antioxidant octadecanoic acid in talh honey while heneicosane and hydrocinnamic acid were present at a higher content in manuka honey. The molecular mechanism was to limit the expression of inflammatory signals, including COX-2 and NF-κB, and the expression of the apoptotic signal, BAX and caspase-3 while inducing Bcl-2 expression.
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Antineoplásicos/efeitos adversos , Cisplatino/toxicidade , Mel/análise , Rim/efeitos dos fármacos , Fígado/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Substâncias Protetoras/administração & dosagem , Proteína X Associada a bcl-2/metabolismo , Animais , Antioxidantes/administração & dosagem , Apoptose/efeitos dos fármacos , Caspase 3/genética , Caspase 3/metabolismo , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Humanos , Rim/metabolismo , Fígado/metabolismo , Masculino , NF-kappa B/genética , NF-kappa B/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Proteína X Associada a bcl-2/genéticaRESUMO
INTRODUCTION: Spinal cord injury (SCI) is a life-disrupting condition in which the first few days are the most critical. Secondary conditions remain the main causes of death for people with SCI. The response of different cell types to SCI and their role at different times in the progression of secondary degeneration are not well understood. The aim of this study was to study the histopathological changes of compressed spinal cord injury (CSCI) in a rat model. MATERIAL AND METHODS: Forty adult male Sprague-Dawley rats were divided into four groups. In group I, the rats were left without any surgical intervention (control). In group II, the rats were subjected to laminectomy without spinal cord compression (sham-operated). In group III, the rats were sacrificed one day after CSCI. In group IV, the rats were sacrificed seven days after CSCI. The light microscopy was employed to study the morphology using H&E, osmic acid staining and immunohistochemistry to detect glial fibrillary acidic protein (GFAP). The electron microscopy was applied for ultrastructure study. RESULTS: Histopathological examination of the posterior funiculus of the white matter revealed minute hemorrhages and localized necrotic areas on day 1, which transformed to areas of cavitation and fibrinoid necrosis surrounded by a demarcating rim of numerous astrocytes by day 7. The mean percentage of area of GFAP expression increased significantly by day 7. Osmic acid staining revealed swollen nerve fibers after one day, while numerous fibers had been lost by day 7. An ultrastructure study revealed swollen redundant thinned myelin and myelin splitting, as well as degeneration of axoplasm on day 1. On day 7, layers of the myelin sheath were folded and wrinkled with partial or complete demyelination areas. The myelin lamellae were disorganized and loose. The G-ratio was significantly greater on day 1 than day 7 after CSCI. CONCLUSIONS: In the rat model of CSCI details of the progressive spinal cord injury can be analyzed by morphological methods and may be helpful in the identification of the onset and type of clinical intervention.
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Compressão da Medula Espinal/patologia , Traumatismos da Medula Espinal/patologia , Medula Espinal/patologia , Medula Espinal/ultraestrutura , Animais , Modelos Animais de Doenças , Imuno-Histoquímica , Masculino , Ratos , Ratos Sprague-Dawley , Fatores de TempoRESUMO
Gastric ulcers are a major problem worldwide with no effective treatment. The objective of this study was to evaluate the use of manuka honey in the treatment of acetic acid-induced chronic gastric ulcers in rats. Different groups of rats were treated with three different concentrations of honey. Stomachs were checked macroscopically for ulcerative lesions in the glandular mucosa and microscopically for histopathological alterations. Treatment with manuka honey significantly reduced the ulcer index and maintained the glycoprotein content. It also reduced the mucosal myeloperoxidase activity, lipid peroxidation (MDA), and the inflammatory cytokines (TNF-α, IL-1ß, and IL-6) as compared to untreated control group. In addition, honey-treated groups showed significant increase in enzymatic (GPx and SOD) and nonenzymatic (GSH) antioxidants besides levels of the anti-inflammatory cytokine IL-10. Flow cytometry studies showed that treatment of animals with manuka honey has normalized cell cycle distribution and significantly lowered apoptosis in gastric mucosa. In conclusion, the results indicated that manuka honey is effective in the treatment of chronic ulcer and preservation of mucosal glycoproteins. Its effects are due to its antioxidant and anti-inflammatory properties that resulted in a significant reduction of the gastric mucosal MDA, TNF-α, IL-1ß, and IL-6 and caused an elevation in IL-10 levels.
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Food and Drug Administration (FDA, USA)-approved category B antibiotics are commonly prescribed to treat infections during pregnancy. The aim of this study was to investigate antibiotic-induced changes in gut microbiota (GM) that occur during pregnancy. The 16S rRNA amplicon deep-sequencing method was used to analyze the effect of category B antibiotics (azithromycin, amoxicillin and cefaclor) on GM during pregnancy using a rat model. The GM composition was substantially modulated by pregnancy and antibiotics administration. Firmicutes, Bacteroidetes, Proteobacteria, Chlamydiae, Actinobacteria, and Cyanobacteria were the dominant phyla. Antibiotic treatment during pregnancy increased the relative abundance of Proteobacteria and reduced Firmicutes. The genera Shigella, Streptococcus, Candidatus Arthromitus, and Helicobacter were significantly (p < 0.05) more abundant during pregnancy. Antibiotics significantly (p < 0.05) reduced the relative abundance of Lactobacillus but increased that of Enterobacter. There was a significant (p < 0.05) decrease in Lactobacillus sp., Lactobacillus gallinarum and Lactobacillus crispatus during pregnancy. Antibiotic treatment reduced bacterial diversity; the lowest number of operational taxonomic units (OTUs) were detected in the cefaclor-treated groups. Antibiotics significantly (p < 0.05) promoted weight gain during pregnancy, and increased relative abundance of Shigella sonnei, Enterococcus hormaechei, and Acinetobacter sp. GM perturbations were accompanied by increases in Proteobacteria abundance and weight gain in pregnancy following antibiotic treatment.
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Gastric ulcers are among the most common diseases affecting humans. This study aimed at investigating the gastroprotective effects of manuka honey against ethanol-induced gastric ulcers in rats. The mechanism by which honey exerts its antiulcer potential was elucidated. Four groups of rats were used: control, ethanol (ulcer), omeprazole, and manuka honey. Stomachs were examined macroscopically for hemorrhagic lesions in the glandular mucosa, histopathological changes, and glycoprotein detection. The effects of oxidative stress were investigated using the following indicators: gastric mucosal nitric oxide (NO), reduced glutathione (GSH), lipid peroxide (MDA, measured as malondialdehyde) glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase. Plasma tumour necrosis factor-α, interleukin-1ß, and IL-6 were also measured. Manuka honey significantly decreased the ulcer index, completely protected the mucosa from lesions, and preserved gastric mucosal glycoprotein. It significantly increased gastric mucosal levels of NO, GSH, GPx, and SOD. Manuka honey also decreased gastric mucosal MDA and plasma TNF-α, IL-1ß, and IL-6 concentrations. In conclusion, manuka honey likely exerted its antiulcer, effect by keeping enzymatic (GPx and SOD) and nonenzymatic (GSH and NO) antioxidants as well as inflammatory cytokines (TNF-α, IL-1ß, and IL-6) in a reduced form, inhibited lipid peroxidation (MDA), and preserved mucous glycoproteins levels.
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Anti-Inflamatórios/uso terapêutico , Antioxidantes/uso terapêutico , Mel , Úlcera Gástrica/tratamento farmacológico , Animais , Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Glicemia/metabolismo , Catalase/metabolismo , Colesterol/sangue , Modelos Animais de Doenças , Comportamento de Ingestão de Líquido/efeitos dos fármacos , Etanol , Comportamento Alimentar/efeitos dos fármacos , Glutationa/metabolismo , Glutationa Peroxidase/metabolismo , Glicoproteínas/metabolismo , Interleucina-1beta/sangue , Interleucina-6/sangue , Masculino , Malondialdeído/metabolismo , Óxido Nítrico/metabolismo , Ratos , Coloração e Rotulagem , Úlcera Gástrica/patologia , Superóxido Dismutase/metabolismo , Triglicerídeos/sangue , Fator de Necrose Tumoral alfa/sangue , Aumento de Peso/efeitos dos fármacosRESUMO
Hepatitis C virus (HCV) is a major health problem worldwide. Early detection of the infection will help better management of the infected cases. The monoclonal antibodies (mAb) of mice are predominantly used for the immunodiagnosis of several viral, bacterial, and parasitic antigens. Serological detection of HCV antigens and antibodies provide simple and rapid methods of detection but lack sensitivity specially in the window phase between the infection and antibody development. Human mAb are used in the immunotherapy of several blood malignancies, such as lymphoma and leukemia, as well as for autoimmune diseases. In this review article, we will discuss methods of mouse and human monoclonal antibody production. We will demonstrate the role of mouse mAb in the detection of HCV antigens as rapid and sensitive immunodiagnostic assays for the detection of HCV, which is a major health problem throughout the world, particularly in Egypt. We will discuss the value of HCV-neutralizing antibodies and their roles in the immunotherapy of HCV infections and in HCV vaccine development. We will also discuss the different mechanisms by which the virus escape the effect of neutralizing mAb. Finally, we will discuss available and new trends to produce antibodies, such as egg yolk-based antibodies (IgY), production in transgenic plants, and the synthetic antibody mimics approach.
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Marine sponges are found to be a rich source of bioactive compounds which show a wide range of biological activities including antiviral, antibacterial, and anti-inflammatory activities. This study aimed to investigate the possible anti-inflammatory, antioxidant and immunomodulator effects of the methanolic extract of the Red Sea marine sponge Xestospongia testudinaria. The chemical composition of the Xestospongia testudinaria methanolic extract was determined using Gas chromatography-mass spectroscopy (GC-MS) analysis. DPPH (2, 2-diphenyl-1-picryl-hydrazyl) was measured to assess the antioxidant activity of the sponge extract. Carrageenan-induced rat hind paw edema was adopted in this study. Six groups of rats were used: group1: Control, group 2: Carrageenan, group 3: indomethacin (10 mg/kg), group 4-6: Xestospongia testudinaria methanolic extract (25, 50, and 100 mg/kg). Evaluation of the anti-inflammatory activity was performed by both calculating the percentage increase in paw weight and hisopathologically. Assessment of the antioxidant and immunomodulatory activity was performed. GC-MS analysis revealed that there were 41 different compounds present in the methanolic extract. Sponge extract exhibited antioxidant activity against DPPH free radicals. Xestospongia testudinaria methanolic extract (100 mg/kg) significantly decreased % increase in paw weight measured at 1, 2, 3 and 4 h after carrageenan injection. Histopathologically, the extract caused a marked decrease in the capillary congestion and inflammatory cells infiltrate. The extract decreased paw malondialdehyde (MDA) and nitric oxide (NO) and increased the reduced glutathione (GSH), glutathione peroxidase (GPx), and catalase (CAT) activity. It also decreased the inflammatory cytokines, tumor necrosis factor-α (TNF-α), interleukin-1 ß(IL-1ß) and IL-6. The results of this study demonstrated the anti-inflammatory, antioxidant, and immunomodulatory effects of the methanolic extract of the Red Sea sponge Xestospongia testudinaria (100 mg/kg).
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Anti-Inflamatórios , Misturas Complexas , Edema/tratamento farmacológico , Sequestradores de Radicais Livres , Fatores Imunológicos , Xestospongia/química , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Misturas Complexas/química , Misturas Complexas/farmacologia , Avaliação Pré-Clínica de Medicamentos , Edema/induzido quimicamente , Edema/patologia , Sequestradores de Radicais Livres/química , Sequestradores de Radicais Livres/farmacologia , Fatores Imunológicos/química , Fatores Imunológicos/farmacologia , Oceano Índico , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Recent studies have demonstrated that marine sponges and their active constituents exhibited several potential medical applications. This study aimed to evaluate the possible hepatoprotective role as well as the antioxidant effect of the Red Sea Suberea mollis sponge extract (SMSE) on carbon tetrachloride- (CCl4-) induced acute liver injury in rats. In vitro antioxidant activity of SMSE was evaluated by 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) assay. Rats were orally administered three different concentrations (100, 200, and 400 mg/kg) of SMSE and silymarin (100 mg/kg) along with CCl4 (1 mL/kg, i.p., every 72 hr) for 14 days. Plasma aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), and total bilirubin were measured. Hepatic malondialdehyde (MDA), reduced glutathione (GSH), nitric oxide (NO), superoxide dismutase (SOD), glutathione peroxidase (GPx), and catalase (CAT) were also measured. Liver specimens were histopathologically examined. SMSE showed strong scavenging activity against free radicals in DPPH assay. SMSE significantly reduced liver enzyme activities. Moreover, SMSE significantly reduced hepatic MDA formation. In addition, SMSE restored GSH, NO, SOD, GPx, and CAT. The histopathological results confirmed these findings. The results of this study suggested a potent protective effect of the SMSE against CCl4-induced hepatic injury. This may be due to its antioxidant and radical scavenging activity.
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The present study tested the hypothesis that prenatal exposure of neonate Outbred albino mice to Schistosoma mansoni antigens (Ags) or antibodies (Abs) modulates their immunity against postnatal responses to infection. Persistence of maternal S. mansoni Abs and/or Ags in mice born to S. mansoni-infected mothers (IF-IMs) and noninfected mothers (IF-NMs) for up to 8 weeks after delivery was investigated. A higher level of anti-S. mansoni IgG Ab was detected in sera of 1-week-old mice born to IF-IM compared to controls. Then, immunoglobulin (Ig)G gradually decreased to the eight week. No anti-S. mansoni IgM Ab was detected in sera of these offspring at any week after delivery. Schistosoma Ags were detected in liver and kidney tissues of mice born to infected mothers. However, Ags decreased markedly till the sixth week in the liver but increased significantly at the sixth week in the kidney. Eight-week-old mice born to infected and noninfected mothers were infected with 200 S. mansoni ceracriae. Their sera and livers were collected for testing IgG and granuloma formation 6 weeks postinfection. Worms were collected via portal perfusion and counted. Anti-S. mansoni IgG level, size and number of liver granuloma, and worm burden were significantly reduced in the offspring of infected mothers. These data suggest that in utero exposure of Outbred albino mice to S. mansoni may attenuate the pathogenesis of S. mansoni in subsequent challenge.