High Levels Of Serum Antibodies Research Articles

Enhancing immune response, antioxidant capacity, and gut health in growing beagles through a chitooligosaccharide diet.

Chitooligosaccharides (COS) have attracted significant attention due to their unique biological activities, water solubility, and absorbable properties. The objective of the present study was to investigate the impact of COS-supplemented diets on the immune response, antioxidative capacity, hematology, serum biochemistry, and modulation of intestinal microbiota in growing beagles. Twelve weaning male beagles (6 weeks old; weighing 3.6 ± 0.6 kg) were fed either a control diet (food without COS, n = 6) or a COS-supplemented diet (n = 6) twice daily for 7 weeks. Blood samples collected at weeks 4 and 7 indicated that hematology and serum biochemistry remained unaffected by COS supplementation. Compared with the control group, the test group showed higher levels of serum antibodies against the canine distemper virus and parvovirus, higher levels of immunoglobulin A, G, and M, and increased activities of superoxide dismutase, glutathione peroxidase, and catalase. In addition, COS was observed to modulate the intestinal flora by enhancing the presence of probiotics, such as Muribaculaceae, Prevotellaceae_Ga6A1_group, Lactobacillus, Collinsella, Blautia, and Lachnospiraceae_NK4A136_group. In summary, a COS-supplemented diet could effectively improve dog health by regulating immune function and antioxidant responses and modulating intestinal microbiota. This study highlights the potentiality of using COS as a valuable nutraceutical for growing dogs.

Evaluation of macular choroidal and microvascular network changes by activity scores and serum antibodies in thyroid eye patients and healthy subjects.

To investigate the choroidal thickness and the microvascular network changes around the macula in thyroid eye disease (TED) patients at different stages and the relationship of those changes with risk factors, serum antibodies and the severity of TED. A total of 85 participants were enrolled. All participants underwent ophthalmology and endocrinology examinations. Subfoveal choroidal thickness (SFCT), superficial (s) and deep (d) foveal avascular zone (FAZ) area, mean (m) and central (c) superficial vascular density (SVD), deep vascular density (DVD) measurements of the enrolled cases were performed with Topcon swept source optical coherence tomography (OCT)/OCT angiography (OCTA) DRI OCT Triton. Multiple linear regression analysis was used to explore the associations between SFCT, FAZ area, SVD, DVD and the relevant factors of TED. Those with active TED patients had higher c-DVD and m-DVD levels (P<0.05), however there is no statistically significant difference in SFCT between active and stable TED patients. Among the serum antibodies, it was observed that s-FAZ and d-FAZ increased, c-SVD and m-SVD decreased in patients with high thyroid stimulating hormone-receptor autoantibodies (TRAB) level, whereas SFCT thickened in patients with high levels of both TRAB and human thyroglobulin (hTG). There was no significant difference in SFCT, FAZ, SVD and DVD measurement at gender, between hyperthyroid and euthyroid patients and among those with or without thyroid papillary carcinoma. The results show that both disease activation and serum antibodies differentially affect both superficial and deep retinal vascular density. It has also been shown that high serum antibody levels affect choroidal thickness independent of clinical activity.

Role of age in dynamics of autoantibodies in pediatric Celiac disease

BackgroundCeliac disease (CD) is characterized by elevated serum titers of autoantibodies IgA anti-tissue transglutaminase 2 (TGA-IgA) and IgA anti-endomysial (EMA), with small bowel mucosa atrophy. We evaluated age differences between CD children exhibiting variable antibody titers at diagnosis.MethodsCD children diagnosed between January 2014 and June 2019, according to 2012 ESPGHAN guidelines were studied. All had EMA and TGA-IgA measurements, while a proportion of them underwent esophagogastroduodenoscopy (EGD). Patients were grouped based on serum TGA-IgA titers normalized to the upper limit of normal (ULN) and differences in median age (years) assessed by analysis of variance (ANOVA) and creation of orthogonal contrasts.ResultsCD was diagnosed in 295 subjects (median age: 4.4 [IQR: 2.60–8.52]) with a biopsy sparing protocol (high titer: ≥ 10xULN) and in 204 by EGD biopsy. Of the latter, 142 (median age: 8.5 [IQR: 5.81–11.06]) and 62 (median age: 9.5 [IQR: 6.26–12.76]) had a low (< 5xULN) and a moderate (≥ 5 < 10xULN) TGA-IgA titer, respectively. Potential CD was diagnosed in 20 patients (median age: 3.6 [IQR: 2.47–6.91]). The median age was significantly lower in the no-biopsy group (ANOVA: F(3, 516) = 25.98, p < .001) than in low- and moderate titer groups (p < 0.0001), while there was no statistical difference between biopsy-sparing and potential CD groups.ConclusionCD patients with greatly elevated antibody titers (≥ 10xULN) were diagnosed at an earlier age than those with lower titers. This may indicate that an increase in TGA-IgA is independent of age and suggests a polarization of autoimmunity in younger individuals with higher serum antibody levels.

A multi-epitope vaccine GILE against Echinococcus Multilocularis infection in mice.

The objective of this study is to construct a multi-epitope vaccine GILE containing B-cell and T-cell epitopes against Echinococcus Multilocularis (E. multilocularis) infection based on the dominant epitopes of E. multilocularis EMY162, LAP, and GLUT1. The structure and hydrophobicity of GILE were predicted by SWISSMODEL, pyMOL, SOPMA and VMD, and its sequence was optimized by Optimum™ Codon. The GILE gene was inserted into pCzn1 and transformed into Escherichia coli Arctic express competent cells. IPTG was added to induce the expression of recombinant proteins. High-purity GILE recombinant protein was obtained by Ni-NTA Resin. BALB/c mice were immunized with GILE mixed with Freund's adjuvant, and the antibody levels and dynamic changes in the serum were detected by ELISA. Lymphocyte proliferation was detected by MTS. The levels of IFN-g and IL-4 were detected by ELISpot and flow cytometry (FCM). T cells were detected by FCM. The growth of hepatic cysts was evaluated by Ultrasound and their weights were measured to evaluate the immune protective effect of GILE. The SWISS-MODEL analysis showed that the optimal model was EMY162 95-104-LAP464-479-LAP495-510-LAP396-410-LAP504-518-EMY162112-126. The SOPMA results showed that there were Alpha helix (14.88%), Extended strand (26.25%), Beta turn (3.73%) and Random coil (45.82%) in the secondary structure of GILE. The restriction enzyme digestion and sequencing results suggested that the plasmid pCzn1-GILE was successfully constructed. The SDSPAGE results indicated that the recombinant protein was 44.68 KD. The ELISA results indicated that mice immunized with GILE showed higher levels of serum antibodies compared to the PBS group. The FCM and ELISpot results indicated that mice immunized with GILE secreted more IFN-g and IL-4. Immunization with GILE also led to a significant decrease in the maximum diameter and weight of cysts and stimulated the production of CD4+ and CD8+ T Cell. A multi-epitope vaccine GILE with good immunogenicity and antigenicity has been successfully constructed in this study, which may provide important theoretical and experimental bases for the prevention and treatment of E. multilocularis infection.

Plant-Produced Mouse-Specific Zona Pellucida 3 Peptide Induces Immune Responses in Mice.

Contraceptive vaccines are designed to stimulate autoimmune responses to molecules involved in the reproductive process. A mouse-specific peptide from zona pellucida 3 (mZP3) has been proposed as a target epitope. Here, we employed a plant expression system for the production of glycosylated mZP3 and evaluated the immunogenicity of plant-produced mZP3-based antigens in a female BALB/c mouse model. In the mZP3-1 antigen, mZP3 fused with a T-cell epitope of tetanus toxoid, a histidine tag, and a SEKDEL sequence. A fusion antigen (GFP-mZP3-1) and a polypeptide antigen containing three repeats of mZP3 (mZP3-3) were also examined. Glycosylation of mZP3 should be achieved by targeting proteins to the endoplasmic reticulum. Agrobacterium-mediated transient expression of antigens resulted in successful production of mZP3 in Nicotiana benthamiana. Compared with mZP3-1, GFP-mZP3-1 and mZP3-3 increased the production of the mZP3 peptide by more than 20 and 25 times, respectively. The glycosylation of the proteins was indicated by their size and their binding to a carbohydrate-binding protein. Both plant-produced GFP-mZP3-1 and mZP3-3 antigens were immunogenic in mice; however, mZP3-3 generated significantly higher levels of serum antibodies against mZP3. Induced antibodies recognized native zona pellucida of wild mouse, and specific binding of antibodies to the oocytes was observed in immunohistochemical studies. Therefore, these preliminary results indicated that the plants can be an efficient system for the production of immunogenic mZP3 peptide, which may affect the fertility of wild mice.

H11-induced immunoprotection is predominantly linked to N-glycan moieties during Haemonchus contortus infection.

Nematodes are one of the largest groups of animals on the planet. Many of them are major pathogens of humans, animals and plants, and cause destructive diseases and socioeconomic losses worldwide. Despite their adverse impacts on human health and agriculture, nematodes can be challenging to control, because anthelmintic treatments do not prevent re-infection, and excessive treatment has led to widespread drug resistance in nematode populations. Indeed, many nematode species of livestock animals have become resistant to almost all classes of anthelmintics used. Most efforts to develop commercial anti-nematode vaccines (native or recombinant) for use in animals and humans have not succeeded, although one effective (dead) vaccine (Barbervax) has been developed to protect animals against one of the most pathogenic parasites of livestock animals - Haemonchus contortus (the barber's pole worm). This vaccine contains native molecules, called H11 and H-Gal-GP, derived from the intestine of this blood-feeding worm. In its native form, H11 alone consistently induces high levels (75-95%) of immunoprotection in animals against disease (haemonchosis), but recombinant forms thereof do not. Here, to test the hypothesis that post-translational modification (glycosylation) of H11 plays a crucial role in achieving such high immunoprotection, we explored the N-glycoproteome and N-glycome of H11 using the high-resolution mass spectrometry and assessed the roles of N-glycosylation in protective immunity against H. contortus. Our results showed conclusively that N-glycan moieties on H11 are the dominant immunogens, which induce high IgG serum antibody levels in immunised animals, and that anti-H11 IgG antibodies can confer specific, passive immunity in naïve animals. This work provides the first detailed account of the relevance and role of protein glycosylation in protective immunity against a parasitic nematode, with important implications for the design of vaccines against metazoan parasites.

Oral vaccination with feed-based live attenuated vaccine protects large yellow croaker against Pseudomonas plecoglossicida infection

Pseudomonas plecoglossicida is a facultative Gram-negative bacteria that causes ‘visceral granulomas disease’ (VGD) in large yellow croaker (LYC; Larimichthys crocea) and several other fish species leading to high economic losses. Currently, there are no commercial vaccines against VGD. Thus, we evaluated the efficacy of a live attenuated vaccine (LAV) made by our laboratory in LYC administered orally in alginate-coated LAV-loaded feeds. To evaluate the tolerance of the LAV in the acidic environment of the stomach, we treated the alginate-coated LAV with a simulated gastric juice without pepsin. There was a 10-fold decrease in the number of viable LAV cells after treatment with the simulated gastric juice (pH 2.0). Despite so, we found a significant increase in LAV cells in the liver and hindgut within 4 h post first feeding of oral vaccine (hpff), which increased to high levels by 24 hpff indicating that LAV cells that survived the stomach acidic environment reached the hindgut for adsorption. Although there was a transient change in the composition of the gut microbiota at 24 hpff, analysis using the Shannon and β-diversity indices showed a similarity in the bacterial composition found before and after vaccination indicating that the LAV had no long-term adverse effect on the gut microbiota. Systemic and mucosal humoral responses were characterized by high serum antibody levels and upregulation of the pig and sign while activation of cell-mediated immunity was characterized by upregulation of MHC-Iα, MHC-IIβ, Prf1 and IFNγ genes. The overall protection in vaccinated LYC was estimated at relative percent survival (RPS) of 60%. Altogether, this study has shown that LAVs can be used as oral vaccines to confer protective immunity without impacting adverse effects on the fish microbiota.

Immune-hormonal imbalance in chemical cancerogenesis

The present article deals with experimental and clinical aspects of immuno-hormonal interactions in chemical carcinogenesis i.e., formation of DNA-adducts with chemical carcinogens as a trigger of tumor initiation; synthesis of specific antibodies as markers of human exposure to environmental carcinogens; immunomodulation of chemical carcinogenesis by the specific antibodies in experimental studies; interactions of antibodies against environmental carcinogens with endogenous steroid hormones in human carcinogenesis; immunological interference and inversion of immuno-hormonal interactions by the action of antibodies against environmental carcinogens; immune stimulation of tumor progression in cancer patients. It is shown that antibodies specific to estradiol and progesterone participate in regulation of serum estradiol and progesterone levels in healthy women. Excessive production of antibodies against benzo[a]pyrene is associated with impaired physiological balance between the levels of antibodies to estradiol and progesterone, thus causing disturbed physiological balance between serum estradiol and progesterone. Immuno-hormonal imbalance promotes tumor initiation, its growth and progression. The new approaches to the personalized cancer immunoprediction and immune prevention are discussed. Coordinated synthesis of antibodies against benzo[a]pyrene and estradiol seems to reflect production of DNA-adducts with genotoxic metabolic effects of these compounds manifesting as synergistic carcinogenic effects upon the target cells. Hence, simultaneously increased levels of serum antibodies against benzo[a]pyrene and estradiol in healthy people may be considered an immunological marker of high oncological risk and an reason to use of new immunoprotective tools against polycyclic aromatic hydrocarbons and phytoestrogens. However, ability of these antibodies to raise the blood serum levels of environmental carcinogens and endogenous estradiol, as shown in vitro and in vivo, excludes the opportunity for active cancer immune prevention. Usage of anticarcinogen vaccines aimed for induction of protective secretory antibodies is likely to further increase high levels of procarcinogenic serum antibodies against benzo[a]pyrene and estradiol, followed by additional enhancement of immuno-hormonal imbalance and promotion of carcinogenesis. Development of probiotics transduced with genes encoding human antibodies against environmental carcinogens may present an alternative approach to cancer immune prevention. The antibodies produced by such probiotics would bind appropriate carcinogens and prevent their invasion into the organism, thus inhibiting emergence of DNA-adducts and suppressing synthesis of specific autoantibodies that may promote carcinogenesis. The aim is to substantiate the concept of immuno-hormonal imbalance for the carcinogen-induced hormone-dependent tumors.

The Correlation between New Serological Markers and Disease Phenotype and Activation in Inflammatory Bowel Disease.

Background: The aim of the study is to assess the correlation between a new antibody panel that is developed against glycans on Crohn's disease (CD) and ulcerative colitis (UC) differentiative diagnosis and disease properties. Methods: In the study, 137 CD and 122 UC patients and 90 controls were included. Anti-saccharomyces cerevisiae IgG (ASCA), anti-laminaribioside IgG (ALCA), anti-chitobioside IgA (ACCA), and anti-mannobioside IgG (AMCA) were tested in serum. Results: While at least 1 of the other 3 serological markers was positive in 89% of ASCA-positive patients, at least 1 of the other 3 serological markers was positive in 77% of ASCA-negative patients. Positivity ratio for a single anticarbohydrate was ALCA 18 (22%), ACCA 5 (12%), and AMCA 16 (23%). A significant correlation was found between ASCA positivity (P<0.001) in operated patients and between ASCA, ALCA, and ACCA positivity (P<0.05) in patients with stricturing and fistulizing CD. According to the ROC analysis, ASCA was found to have the highest area under the curve (0.70-0.82) (correlation coefficient interval 95%). A significant correlation was found between ASCA, ALCA, and ACCA positivity and high serum antibody levels and disease activation (P<0.05). Conclusion: ASCA, ALCA, and ACCA were found to be correlated with the disease complication and activation in CD. ASCA and ALCA were determined as the best markers in the differentiation between CD and UC.

ESPGHAN Position Paper on Management and Follow-up of Children and Adolescents With Celiac Disease.

To gather the current evidence and to offer recommendations for follow-up and management. The Special Interest Group on Celiac Diseases of the European Society of Paediatric Gastroenterology Hepatology and Nutrition formulated ten questions considered to be essential for follow-up care. A literature search (January 2010-March 2020) was performed in PubMed or Medline. Relevant publications were identified and potentially eligible studies were assessed. Statements and recommendations were developed and discussed by all coauthors. Recommendations were voted upon: joint agreement was set as at least 85%. Publications (n = 2775) were identified and 164 were included. Using evidence or expert opinion, 37 recommendations were formulated on: The need to perform follow-up, its frequency and what should be assessed, how to assess adherence to the gluten-free diet, when to expect catch-up growth, how to treat anemia, how to approach persistent high serum levels of antibodies against tissue-transglutaminase, the indication to perform biopsies, assessment of quality of life, management of children with unclear diagnosis for which a gluten-challenge is indicated, children with associated type 1 diabetes or IgA deficiency, cases of potential celiac disease, which professionals should perform follow-up, how to improve the communication to patients and their parents/caregivers and transition from pediatric to adult health care. We offer recommendations to improve follow-up of children and adolescents with celiac disease and highlight gaps that should be investigated to further improve management.

Immunogenicity of an inactivated novel goose parvovirus vaccine for short beak and dwarfism syndrome in Cherry Valley ducks.

Duck short beak and dwarfism syndrome (SBDS) is a viral infectious disease caused by novel goose parvovirus (NGPV), which has been responsible for serious economic losses to the Chinese duck industry in recent years. Currently, there is no effective vaccine against this disease. In this study, we developed an inactivated virus vaccine candidate for SBDS based on NGPV strain DS15 isolated from a duck in China. Immune efficacy was evaluated in 112 ducks, which were randomly divided into vaccination, challenge-control, vaccination-challenge, and blank control groups (28 per group). Clinical characteristics, antibodies, virus excretion, viremia, and pathological changes were monitored. No morbidity or death was observed in the immunized ducks, which showed normal weight and a good mental state. High levels of serum antibodies (optical density at 450nm of ~0.63) were detected in ducks immunized with the inactivated vaccine at 7days post-vaccination (dpv), and the titer of virus-neutralizing antibodies increased from 1:23 to 1:28.5 from 7 to 42dpv. Measurement of the viral load in anal swab, serum, and tissue samples showed that vaccination significantly inhibited the replication of NGPV in immunized ducks. Moreover, NGPV could not be isolated from the spleens of immunized or vaccinated and challenged ducks. Collectively, these results demonstrate that the newly developed inactivated NGPV vaccine, administered in an oil emulsion adjuvant, possesses good immunogenicity and represents a potentially powerful tool for SBDS prevention and control.

Jieduquyuziyin Prescription alleviates hepatic gluconeogenesis via PI3K/Akt/PGC-1α pathway in glucocorticoid-induced MRL/lpr mice

Ethnopharmacological relevanceJieduquyuziyin prescription (JP) is a traditional Chinese medicine (TCM) formula. According to both TCM theory and more than a decade of clinical practice, JP has been testified to be effective for systemic lupus erythematosus (SLE) treatment as an approved hospital prescription in China. Aim of the studyTo determine the effect of JP on the treatment of SLE by glucocorticoid (GC) and to further examine the molecular mechanisms. Materials and methodsWe conducted in vivo experiments to estimate the effect of JP on hepatic gluconeogenesis in MRL/lpr mice treated with GC. Additionally, isoproterenol (ISO) induced hepatic gluconeogenesis model and GC-treated MRL/lpr mouse hepatocytes were carried out in vitro experiments to verify the effect of JP on gluconeogenesis. ResultsThe results showed that JP combined with GC could effectively alleviate the lupus symptoms in MRL/lpr mice and improve the pathological changes of the kidney and liver. And the combination of JP reduced the side effects caused by GC, which was related to the inhibition of GC-induced hepatic gluconeogenesis in MRL/lpr mice. Specifically, JP up-regulated the expression of glucocorticoid receptor (GR) α, phosphoinositide-3-kinase (PI3K) and Akt restrained by GC to reduce the production of forkhead box O1 (FoxO1), peroxisome proliferator-activated receptor-gamma coactivator-1α (PGC-1α), and the gluconeogenic genes phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase). In vivo, the use of JP either alone or with GC could reduce spleen enlargement, high levels of serum antibodies, aggravated urine protein and renal pathological damage in MRL/lpr mice. Furthermore, the glucose content was reduced in the liver of MRL/lpr mice treated with JP, and the liver damage and steatosis were also alleviated. In vitro, the expressions of PI3K and Akt increased and the expressions of FoxO1, PGC-1α, PEPCK and G6Pase decreased after JP treatment in ISO-treated hepatocytes. Compared with MRL/MP mice, we found that JP could significantly inhibit the expression of gluconeogenesis in the hepatocytes of MRL/lpr mice induced by GC to a greater extent. ConclusionsThe therapeutic effect of JP on GC-induced is likely related to hepatic gluconeogenesis, which provides a new perspective to reveal the positive role of JP in SLE.

Assessment of the Feasibility of Using Noninvasive Wearable Biometric Monitoring Sensors to Detect Influenza and the Common Cold Before Symptom Onset

Currently, there are no presymptomatic screening methods to identify individuals infected with a respiratory virus to prevent disease spread and to predict their trajectory for resource allocation. To evaluate the feasibility of using noninvasive, wrist-worn wearable biometric monitoring sensors to detect presymptomatic viral infection after exposure and predict infection severity in patients exposed to H1N1 influenza or human rhinovirus. The cohort H1N1 viral challenge study was conducted during 2018; data were collected from September 11, 2017, to May 4, 2018. The cohort rhinovirus challenge study was conducted during 2015; data were collected from September 14 to 21, 2015. A total of 39 adult participants were recruited for the H1N1 challenge study, and 24 adult participants were recruited for the rhinovirus challenge study. Exclusion criteria for both challenges included chronic respiratory illness and high levels of serum antibodies. Participants in the H1N1 challenge study were isolated in a clinic for a minimum of 8 days after inoculation. The rhinovirus challenge took place on a college campus, and participants were not isolated. Participants in the H1N1 challenge study were inoculated via intranasal drops of diluted influenza A/California/03/09 (H1N1) virus with a mean count of 106 using the median tissue culture infectious dose (TCID50) assay. Participants in the rhinovirus challenge study were inoculated via intranasal drops of diluted human rhinovirus strain type 16 with a count of 100 using the TCID50 assay. The primary outcome measures included cross-validated performance metrics of random forest models to screen for presymptomatic infection and predict infection severity, including accuracy, precision, sensitivity, specificity, F1 score, and area under the receiver operating characteristic curve (AUC). A total of 31 participants with H1N1 (24 men [77.4%]; mean [SD] age, 34.7 [12.3] years) and 18 participants with rhinovirus (11 men [61.1%]; mean [SD] age, 21.7 [3.1] years) were included in the analysis after data preprocessing. Separate H1N1 and rhinovirus detection models, using only data on wearble devices as input, were able to distinguish between infection and noninfection with accuracies of up to 92% for H1N1 (90% precision, 90% sensitivity, 93% specificity, and 90% F1 score, 0.85 [95% CI, 0.70-1.00] AUC) and 88% for rhinovirus (100% precision, 78% sensitivity, 100% specificity, 88% F1 score, and 0.96 [95% CI, 0.85-1.00] AUC). The infection severity prediction model was able to distinguish between mild and moderate infection 24 hours prior to symptom onset with an accuracy of 90% for H1N1 (88% precision, 88% sensitivity, 92% specificity, 88% F1 score, and 0.88 [95% CI, 0.72-1.00] AUC) and 89% for rhinovirus (100% precision, 75% sensitivity, 100% specificity, 86% F1 score, and 0.95 [95% CI, 0.79-1.00] AUC). This cohort study suggests that the use of a noninvasive, wrist-worn wearable device to predict an individual's response to viral exposure prior to symptoms is feasible. Harnessing this technology would support early interventions to limit presymptomatic spread of viral respiratory infections, which is timely in the era of COVID-19.

Long‐term follow‐up of convalescent pigs and their offspring after an outbreak of acute African swine fever in Vietnam

African swine fever (ASF) is a contagious haemorrhagic disease in pigs and has become endemic in several Vietnam provinces since the first outbreak in 2019. The presence of carriers and the recurrence of disease in the surviving swine herd after an ASF outbreak has not previously been properly evaluated. In this study, pigs naturally infected with an acute form of ASF were allowed to recover from the disease. A serological follow-up was conducted for more than 14 months with 14 convalescent gilts and their offspring. All convalescent animals had long lasting high serum antibody levels without persistent viremia. They also did not excrete virus via nasal discharge post-recovery. These convalescent pigs could partially perform as replacement gilts despite the fact that ASF affected reproductive performance. Here, we confirmed that there were neither the carriers of nor recurrence of disease in the convalescent pigs and their offspring following the outbreak of acute ASF. These findings may facilitate efforts to design a new farming model in ASF endemic provinces in Vietnam where there is a lack of a repopulation strategy due to the limited funding received from the local regulatory authorities.

P-NE013. Distal myasthenia gravis – A missed clinical diagnosis

Introduction . What is well known: Myasthenia gravis presents with ocular, bulbar and proximal limb weakness. What is less known: Myasthenia gravis can rarely present with predominant distal weakness of hand muscles. Myasthenia gravis (MG) is an immune mediated, post-synaptic neuromuscular transmission disorder, presenting with predominant ocular, bulbar and/or proximal fluctuating weakness and fatiguability. Distal and asymmetric onset of symptoms in MG is rare and could lead to delay in diagnosis especially when fatiguability or fluctuation is not clearly reported as a prominent initial symptom - as seen in our patients. Results . We report 3 cases of distal MG who presented with predominant distal asymmetric hand weakness. Their clinically diagnoses for referral to the electrodiagnostic (EDx) laboratory were chronic inflammatory demyelinating polyneuropathy, C8T1 motor radiculopathy and multifocal motor neuropathy. It was in the EDx laboratory that the diagnosis of MG was suspected in these patients. Since the routine EDx tests were normal in all 3 patients - a repetitive nerve stimulation test (RNST) was done - which was positive in the affected muscles and hence a diagnosis of distal MG made (This is the protocol followed in our laboratory). These patients were then found to have high serum levels of antibodies to acetylcholine receptors - confirming the diagnosis of MG. RNST is not routinely done in all patients sent for electrodiagnostic testing and none of these patients had a reference for the same – however using a protocol based method of EDx testing, leading to the diagnosis of a treatable condition. Conclusion . Electrodiagnosticians need to be aware of this presentation of MG and follow protocols or else the diagnosis could be missed.

A Vaccine Based on Kunitz-Type Molecule Confers Protection Against Fasciola hepatica Challenge by Inducing IFN-γ and Antibody Immune Responses Through IL-17A Production.

Fasciola hepatica is helminth parasite found around the world that causes fasciolosis, a chronic disease affecting mainly cattle, sheep, and occasionally humans. Triclabendazole is the drug of choice to treat this parasite. However, the continuous use of this drug has led to the development of parasite resistance and, consequently, the limitation of its effectiveness. Hence, vaccination appears as an attractive option to develop. In this work, we evaluated the potential of F. hepatica Kunitz-type molecule (FhKTM) as an antigen formulated with a liquid crystal nanostructure formed by self-assembly of 6-O-ascorbyl palmitate ester (Coa-ASC16) and the synthetic oligodeoxynucleotide containing unmethylated cytosine-guanine motifs (CpG-ODN) during an experimental model of fasciolosis in mice, and we further dissected the immune response associated with host protection. Our results showed that immunization of mice with FhKTM/CpG-ODN/Coa-ASC16 induces protection against F. hepatica challenge by preventing liver damage and improving survival after F. hepatica infection. FhKTM/CpG-ODN/Coa-ASC16-immunized mice elicited potent IFN-γ and IL-17A with high levels of antigen-specific IgG1, IgG2a, and IgA serum antibodies. Strikingly, IL-17A blockade during infection decreased IgG2a and IgA antibody levels as well as IFN-γ production, leading to an increase in mortality of vaccinated mice. The present study highlights the potential of a new vaccine formulation to improve control and help the eradication of F. hepatica infection, with potential applications for natural hosts such as cattle and sheep.

Increasing of SIgA serum levels may reflect subclinical intestinal involvement in non-radiographic axial and peripheral spondyloarthritis.

The evidence shows that previous infection with enteric pathogens is a requirement to develop pSpA. Based on our previous results, variances on regulation of SIgA might influence SpA activity; thus, the aim of this study was to correlate the levels of SIgA, IgA against some enteric bacteria, and IL-17, IL-21, and IL-6 with clinical features in a group of SpA patients. Twenty-six pSpA, 20 nr-axSpA, 60 healthy volunteers (HV), and 34 patients with inflammatory bowel diseases (IBD) were included. All subjects were assessed to measure SIgA, total and specific IgA for enteric bacteria, and IL-17, IL-21, and IL-6 levels and clinical variables. For SpA patients, the diagnosis was verified 5 years after first evaluation to assess the risk of developing r-axSpA. SIgA levels were significantly higher in SpA patients than in HV and IBD (p < 0.0001 and p = 0.047, respectively). However, no differences for SIgA neither total IgA were found among the SpAsubtypes (p = 0.624). Only IL-6 was higher in SpA than HV (p = 0.013). An inverse correlation was demonstrated for SIgA and BASFI (r: - 0.45; p = 0.003), BASDAI (r: - 0.39; p = 0.0123), ASDAS-CRP (r: - 0.37; p = 0.014), and ASDAS-ESR (r: - 0.45; p = 0.0021). There was no evidence of risk of developing r-axSpA in patients who previously showed high levels of serum antibodies. The results show that pSpA as well as nr-axSpA share a similar SIgA-intestinal involvement independently of a previous infection. This suggests that serum SIgA increases are evidence of subclinical intestinal compromise which could have influence on disease activity but not in this progression. Key Point • The levels of SIgA, IgA against some enteric bacteria, and IL-17, IL-21, and IL-6 are correlated with clinical features in a group of SpA patients.

A plant-based transient expression system for the rapid production of highly immunogenic Hepatitis E virus-like particles.

Hepatitis E virus (HEV) infection is a major cause of acute hepatitis worldwide. The aim of the study is the development of plant expression system for the production of virus-like particles formed by HEV capsid and the characterization of their immunogenicity. Open reading frame (ORF) 2 encodes the viral capsid protein and possesses candidate for vaccine production. In this study, we used truncated genotype 3 HEV ORF 2 consisting of aa residues 110 to 610. The recombinant protein was expressed in Nicotiana benthamiana plants using the self-replicating potato virus X-based vector pEff up to 10% of the soluble protein fraction. The yield of HEV 110-610 after purification was 150-200µg per 1g of green leaf biomass. The recombinant protein formed nanosized virus-like particles. The immunization of mice with plant-produced HEV 110-610 protein induced high levels of HEV-specific serum antibodies. HEV ORF 2 (110-610 aa) can be used as candidate for the development of a plant-produced vaccine against Hepatitis E.

Development of a live vector vaccine against infectious pancreatic necrosis virus in rainbow trout

Infectious pancreatic necrosis virus (IPNV) causes severe viral disease in salmonids, with major economic losses in the global rainbow trout aquaculture industry following from a high mortality rate. In the present study, we aimed to develop a safe and efficient vaccine to protect rainbow trout against IPNV infection. Administered via the immersion route, a live vector vaccine containing the coding region of the IPNV VP2 induced protective immune responses in rainbow trout. For best protection, juvenile rainbow trout should be immunized with a 1:100 dilution of the titer with 1 × 1010.0 ml−1 TCID50 of the live vector vaccine, via 10-min immersion. Expression of the IPNV VP2 gene was confirmed in spleens of vaccinated rainbow trout, and reached the highest level at 3 days post-vaccination (dpv) and thereafter gradually decreased between 3 and 15 dpv. The expression of TLR-3, TLR-7 and TLR-8 was upregulated after vaccination, and the highest expression levels of IFN-1, Mx-1, Mx-3, Vig-1 and Vig-2 were detected at 3 dpv. Four markers of the adaptive immune response (CD4, CD8, IgM and IgT) were continuously increased in the spleens of vaccinated fish as compared with controls, between 3 and 15 dpv. The cumulative percentage mortality differed significantly between the vaccinated fish and the controls (empty-plasmid-vaccinated). The results of the IPNV challenge showed that the live vector vaccine had a high protection rate against IPNV (relative percent survival value of 88.24), and the vaccinated rainbow trout displayed high levels of serum antibodies against IPNV infection. Taken together, our results demonstrate that this is a promising live vector vaccine that could be used to protect rainbow trout against IPNV infection.

Plant-Produced Recombinant Influenza A Virus Candidate Vaccine Based on Flagellin Linked to Conservative Fragments of M2 Protein and Hemagglutintin.

The development of recombinant influenza vaccines with broad spectrum protection is an important task. The combination of conservative viral antigens, such as M2e, the extracellular domain of the transmembrane protein M2, and conserved regions of the second subunit of hemagglutinin (HA), provides an opportunity for the development of universal influenza vaccines. Immunogenicity of the antigens could be enhanced by fusion to bacterial flagellin, the ligand for Toll-like receptor 5, acting as a powerful mucosal adjuvant. In this study, we report the transient expression in plants of a recombinant protein comprising flagellin of Salmonella typhimurium fused to the conserved region of the second subunit of HA (76–130 a.a.) of the first phylogenetic group of influenza A viruses and four tandem copies of the M2e peptide. The hybrid protein was expressed in Nicotiana benthamiana plants using the self-replicating potato virus X-based vector pEff up to 300 µg/g of fresh leaf tissue. The intranasal immunization of mice with purified fusion protein induced high levels of M2e-specific serum antibodies and provided protection against lethal challenge with influenza A virus strain A/Aichi/2/68(H3N2). Our results show that M2e and hemagglutinin-derived peptide can be used as important targets for the development of a plant-produced vaccine against influenza.