scholarly journals Humans surviving cholera develop antibodies against Vibrio cholerae O-specific polysaccharide that inhibit pathogen motility

2020 ◽  
Author(s):  
Richelle C. Charles ◽  
Meagan Kelly ◽  
Jenny M. Tam ◽  
Aklima Akter ◽  
Motaher Hossain ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Vibrio Cholerae ◽  
Polyclonal Antibody ◽  
High Speed ◽  
Polyclonal Antibodies ◽  
Intestinal Lumen ◽  
Dependent Manner ◽  
Specific Antibodies ◽  
Specific Polysaccharide

ABSTRACTThe mechanism of protection against cholera afforded by previous illness or vaccination is currently unknown. We have recently shown that antibodies targeting O-specific polysaccharide (OSP) of Vibrio cholerae correlate highly with protection against cholera. V. cholerae is highly motile and possesses a flagellum sheathed in O-specific polysaccharide (OSP), and motility of V. cholerae correlates with virulence. Using high speed video microscopy, and building upon previous animal-related work, we demonstrate that sera, polyclonal antibody fractions, and OSP-specific monoclonal antibodies recovered from humans surviving cholera block V. cholerae motility at both subagglutinating and agglutinating concentrations. This anti-motility effect is reversed by pre-adsorbing sera and polyclonal antibody fractions with purified OSP; and is associated with OSP-specific but not flagellin-specific monoclonal antibodies. F[ab] fragments of OSP-specific polyclonal antibodies do not inhibit motility, suggesting a requirement for antibody-mediated crosslinking in motility inhibition. We show that OSP-specific antibodies do not directly affect V. cholerae viability, but that OSP-specific monoclonal antibody highly protects against death in the murine cholera model. We used in vivo competitive index studies to demonstrate that OSP-specific antibodies impede colonization and survival of V. cholerae in intestinal tissues, and that this impact is motility-dependent. Our findings suggest that the impedance of motility by antibodies targeting V. cholerae OSP contributes to protection against cholera.IMPORTANCECholera is a severe dehydrating illness of humans caused by Vibrio cholerae. V. cholerae is a highly motile bacterium that has a single flagellum covered in lipopolysaccharide (LPS) displaying O-specific polysaccharide (OSP), and V. cholerae motility correlates with its ability to cause disease. The mechanisms of protection against cholera are not well understood; however, since V. cholerae is a non-invasive intestinal pathogen, it is likely that antibodies that bind the pathogen or its products in the intestinal lumen contribute to protection from infection. Here, we demonstrate that OSP-specific antibodies isolated from humans surviving cholera in Bangladesh inhibit V. cholerae motility and are associated with protection against challenge in a motility-dependent manner.

scholarly journals Humans Surviving Cholera Develop Antibodies against Vibrio cholerae O-Specific Polysaccharide That Inhibit Pathogen Motility

mBio ◽  
2020 ◽  
Vol 11 (6) ◽  
Author(s):  
Richelle C. Charles ◽  
Meagan Kelly ◽  
Jenny M. Tam ◽  
Aklima Akter ◽  
Motaher Hossain ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Vibrio Cholerae ◽  
Polyclonal Antibody ◽  
High Speed ◽  
Intestinal Lumen ◽  
Dependent Manner ◽  
Content Type ◽  
Specific Antibodies ◽  
Specific Polysaccharide

ABSTRACT The mechanism of protection against cholera afforded by previous illness or vaccination is currently unknown. We have recently shown that antibodies targeting O-specific polysaccharide (OSP) of Vibrio cholerae correlate highly with protection against cholera. V. cholerae is highly motile and possesses a flagellum sheathed in OSP, and motility of V. cholerae correlates with virulence. Using high-speed video microscopy and building upon previous animal-related work, we demonstrate that sera, polyclonal antibody fractions, and OSP-specific monoclonal antibodies recovered from humans surviving cholera block V. cholerae motility at both subagglutinating and agglutinating concentrations. This antimotility effect is reversed by preadsorbing sera and polyclonal antibody fractions with purified OSP and is associated with OSP-specific but not flagellin-specific monoclonal antibodies. Fab fragments of OSP-specific polyclonal antibodies do not inhibit motility, suggesting a requirement for antibody-mediated cross-linking in motility inhibition. We show that OSP-specific antibodies do not directly affect V. cholerae viability, but that OSP-specific monoclonal antibody highly protects against death in the murine cholera model. We used in vivo competitive index studies to demonstrate that OSP-specific antibodies impede colonization and survival of V. cholerae in intestinal tissues and that this impact is motility dependent. Our findings suggest that the impedance of motility by antibodies targeting V. cholerae OSP contributes to protection against cholera. IMPORTANCE Cholera is a severe dehydrating illness of humans caused by Vibrio cholerae. V. cholerae is a highly motile bacterium that has a single flagellum covered in lipopolysaccharide (LPS) displaying O-specific polysaccharide (OSP), and V. cholerae motility correlates with its ability to cause disease. The mechanisms of protection against cholera are not well understood; however, since V. cholerae is a noninvasive intestinal pathogen, it is likely that antibodies that bind the pathogen or its products in the intestinal lumen contribute to protection from infection. Here, we demonstrate that OSP-specific antibodies isolated from humans surviving cholera in Bangladesh inhibit V. cholerae motility and are associated with protection against challenge in a motility-dependent manner.

scholarly journals Generation and Characterization of Human Monoclonal Antibodies Targeting Anthrax Protective Antigen following Vaccination with a Recombinant Protective Antigen Vaccine

2015 ◽  
Vol 22 (5) ◽  
pp. 553-560 ◽  
Author(s):  
Xiangyang Chi ◽  
Jianmin Li ◽  
Weicen Liu ◽  
Xiaolin Wang ◽  
Kexin Yin ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Protective Antigen ◽  
Synergistic Effects ◽  
Central Component ◽  
Dependent Manner ◽  
Human Monoclonal Antibodies ◽  
Specific Antibodies ◽  
Anthrax Protective Antigen

ABSTRACTThe anthrax protective antigen (PA) is the central component of the three-part anthrax toxin, and it is the primary immunogenic component in the approved AVA anthrax vaccine and the “next-generation” recombinant PA (rPA) anthrax vaccines. Animal models have indicated that PA-specific antibodies (AB) are sufficient to protect against infection withBacillus anthracis. In this study, we investigated the PA domain specificity, affinity, mechanisms of neutralization, and synergistic effects of PA-specific antibodies from a single donor following vaccination with the rPA vaccine. Antibody-secreting cells were isolated 7 days after the donor received a boost vaccination, and 34 fully human monoclonal antibodies (hMAb) were identified. Clones 8H6, 4A3, and 22F1 were able to neutralize lethal toxin (LeTx) bothin vitroandin vivo. Clone 8H6 neutralized LeTx by preventing furin cleavage of PA in a dose-dependent manner. Clone 4A3 enhanced degradation of nicked PA, thereby interfering with PA oligomerization. The mechanism of 22F1 is still unclear. A fourth clone, 2A6, that was protective onlyin vitrowas found to be neutralizingin vivoin combination with a toxin-enhancing antibody, 8A7, which binds to domain 3 of PA and PA oligomers. These results provide novel insights into the antibody response elicited by the rPA vaccine and may be useful for PA-based vaccine and immunotherapeutic cocktail design.

scholarly journals Murine monoclonal antibodies against RBD of SARS-CoV-2 neutralize authentic wild type SARS-CoV-2 as well as B.1.1.7 and B.1.351 viruses and protect in vivo in a mouse model in a neutralization dependent manner

2021 ◽  
Author(s):  
Fatima Amanat ◽  
Shirin Strohmeier ◽  
Wen-Hsin Lee ◽  
Sandhya Bangaru ◽  
Andrew B Ward ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Severe Acute Respiratory Syndrome ◽  
Mouse Model ◽  
Neutralizing Antibodies ◽  
Dependent Manner ◽  
Receptor Binding Domain ◽  
Wild Type ◽  
Murine Monoclonal Antibodies

After first emerging in December 2019 in China, severe acute respiratory syndrome 2 (SARS-CoV-2) has since caused a pandemic leading to millions of infections and deaths worldwide. Vaccines have been developed and authorized but supply of these vaccines is currently limited. With new variants of the virus now emerging and spreading globally, it is essential to develop therapeutics that are broadly protective and bind conserved epitopes in the receptor binding domain (RBD) or the whole spike of SARS-CoV-2. In this study, we have generated mouse monoclonal antibodies (mAbs) against different epitopes on the RBD and assessed binding and neutralization against authentic SARS-CoV-2. We have demonstrated that antibodies with neutralizing activity, but not non-neutralizing antibodies, lower viral titers in the lungs when administered in a prophylactic setting in vivo in a mouse challenge model. In addition, most of the mAbs cross-neutralize the B.1.351 as well as the B.1.1.7 variants in vitro.

scholarly journals The Ability of Zika virus Intravenous Immunoglobulin to Protect From or Enhance Zika Virus Disease

2021 ◽  
Vol 12 ◽  
Author(s):  
Amelia K. Pinto ◽  
Mariah Hassert ◽  
Xiaobing Han ◽  
Douglas Barker ◽  
Trevor Carnelley ◽  
...  
Keyword(s):  
Virus Infection ◽  
Polyclonal Antibody ◽  
Zika Virus ◽  
Virus Disease ◽  
Polyclonal Antibodies ◽  
Antibody Therapeutics ◽  
The World ◽  
Flavivirus Infection

The closely related flaviviruses, dengue and Zika, cause significant human disease throughout the world. While cross-reactive antibodies have been demonstrated to have the capacity to potentiate disease or mediate protection during flavivirus infection, the mechanisms responsible for this dichotomy are still poorly understood. To understand how the human polyclonal antibody response can protect against, and potentiate the disease in the context of dengue and Zika virus infection we used intravenous hyperimmunoglobulin (IVIG) preparations in a mouse model of the disease. Three IVIGs (ZIKV-IG, Control-Ig and Gamunex®) were evaluated for their ability to neutralize and/or enhance Zika, dengue 2 and 3 viruses in vitro. The balance between virus neutralization and enhancement provided by the in vitro neutralization data was used to predict the IVIG concentrations which could protect or enhance Zika, and dengue 2 disease in vivo. Using this approach, we were able to define the unique in vivo dynamics of complex polyclonal antibodies, allowing for both enhancement and protection from flavivirus infection. Our results provide a novel understanding of how polyclonal antibodies interact with viruses with implications for the use of polyclonal antibody therapeutics and the development and evaluation of the next generation flavivirus vaccines.

scholarly journals Rational design of peptides for identification of linear epitopes and generation of neutralizing monoclonal antibodies against DKK2 for cancer therapy

2020 ◽  
Vol 3 (2) ◽  
pp. 63-70 ◽  
Author(s):  
Rongqing Zhao ◽  
Qian Xiao ◽  
Maohua Li ◽  
Wenlin Ren ◽  
Chenxi Xia ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Tumor Growth ◽  
Wnt Signaling ◽  
Rational Design ◽  
Polyclonal Antibodies ◽  
Wnt Signaling Pathway ◽  
Keyhole Limpet Hemocyanin ◽  
Linear Epitopes

Abstract Dickkopf-related protein 2 (DKK2)is a member of the Dickkopf family in Wnt signaling pathway. Recently, we found that antibodies against DKK2 could activate natural killer (NK) and CD8+ T cells in tumors and inhibit tumor growth. In this paper, we report the rational design of peptides for identification of linear epitopes and generation of neutralizing monoclonal anti-DKK2 antibodies. To break the immune tolerance, we designed and chemically synthesized six peptides corresponding to different regions of DKK2 as immunogens and found five of them could generate mouse polyclonal antibodies that can bind to the active recombinant human DKK2 protein. Neutralizing mouse monoclonal antibodies (5F8 and 1A10) against human DKK2 were successfully developed by immunizing the mice with two different peptides (34KLNSIKSSL42 and 240KVWKDATYS248) conjugated to Keyhole limpet hemocyanin (KLH). The monoclonal antibodies not only abolish DKK2’s suppression of Wnt signaling in vitro but also inhibits tumor growth in vivo. Currently, those two mAbs are undergoing humanization as immunotherapy candidates and may offer a new drug for treatment of human cancers.

scholarly journals Endothelial Claudin

1999 ◽  
Vol 147 (1) ◽  
pp. 185-194 ◽  
Author(s):  
Kazumasa Morita ◽  
Hiroyuki Sasaki ◽  
Mikio Furuse ◽  
Shoichiro Tsukita
Keyword(s):  
Endothelial Cells ◽  
Epithelial Cells ◽  
Blood Vessels ◽  
Polyclonal Antibody ◽  
Polyclonal Antibodies ◽  
Transmembrane Protein ◽  
Velo Cardio Facial Syndrome ◽  
The Brain ◽  
Specific Polyclonal Antibody

Tight junctions (TJs) in endothelial cells are thought to determine vascular permeability. Recently, claudin-1 to -15 were identified as major components of TJ strands. Among these, claudin-5 (also called transmembrane protein deleted in velo-cardio-facial syndrome [TMVCF]) was expressed ubiquitously, even in organs lacking epithelial tissues, suggesting the possible involvement of this claudin species in endothelial TJs. We then obtained a claudin-6–specific polyclonal antibody and a polyclonal antibody that recognized both claudin-5/TMVCF and claudin-6. In the brain and lung, immunofluorescence microscopy with these polyclonal antibodies showed that claudin-5/TMVCF was exclusively concentrated at cell–cell borders of endothelial cells of all segments of blood vessels, but not at those of epithelial cells. Immunoreplica electron microscopy revealed that claudin-5/TMVCF was a component of TJ strands. In contrast, in the kidney, the claudin-5/TMVCF signal was restricted to endothelial cells of arteries, but was undetectable in those of veins and capillaries. In addition, in all other tissues we examined, claudin-5/TMVCF was specifically detected in endothelial cells of some segments of blood vessels, but not in epithelial cells. Furthermore, when claudin-5/TMVCF cDNA was introduced into mouse L fibroblasts, TJ strands were reconstituted that resembled those in endothelial cells in vivo, i.e., the extracellular face–associated TJs. These findings indicated that claudin-5/TMVCF is an endothelial cell–specific component of TJ strands.

scholarly journals Synthetic Fragments of Vibrio cholerae O1 Inaba O-Specific Polysaccharide Bound to a Protein Carrier Are Immunogenic in Mice but Do Not Induce Protective Antibodies

2004 ◽  
Vol 72 (7) ◽  
pp. 4090-4101 ◽  
Author(s):  
Michael D. Meeks ◽  
Rina Saksena ◽  
Xingquan Ma ◽  
Terri K. Wade ◽  
Ronald K. Taylor ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Cell Epitope ◽  
Immunoglobulin M ◽  
Hydroxyl Group ◽  
Enzyme Linked Immunosorbent Assay ◽  
Protective Antibody ◽  
Infant Mouse ◽  
Molar Ratios ◽  
Specific Polysaccharide

ABSTRACT Development of Vibrio cholerae lipopolysaccharide (LPS) as a cholera vaccine immunogen is justified by the correlation of vibriocidal anti-LPS response with immunity. Two V. cholerae O1 LPS serotypes, Inaba and Ogawa, are associated with endemic and pandemic cholera. Both serotypes induce protective antibody following infection or vaccination. Structurally, the LPSs that define the serotypes are identical except for the terminal perosamine moiety, which has a methoxyl group at position 2 in Ogawa but a hydroxyl group in Inaba. The terminal sugar of the Ogawa LPS is a protective B-cell epitope. We chemically synthesized the terminal hexasaccharides of V. cholerae serotype Ogawa, which comprises in part the O-specific polysaccharide component of the native LPS, and coupled the oligosaccharide at different molar ratios to bovine serum albumin (BSA). Our initial studies with Ogawa immunogens showed that the conjugates induced protective antibody. We hypothesized that antibodies specific for the terminal sugar of Inaba LPS would also be protective. Neoglycoconjugates were prepared from synthetic Inaba oligosaccharides (disaccharide, tetrasaccharide, and hexasaccharide) and BSA at different levels of substitution. BALB/c mice responded to the Inaba carbohydrate (CHO)-BSA conjugates with levels of serum antibodies of comparable magnitude to those of mice immunized with Ogawa CHO-BSA conjugates, but the Inaba-specific antibodies (immunoglobulin M [IgM] and IgG1) were neither vibriocidal nor protective in the infant mouse cholera model. We hypothesize that the anti-Inaba antibodies induced by the Inaba CHO-BSA conjugates have enough affinity to be screened via enzyme-linked immunosorbent assay but not enough to be protective in vivo.

scholarly journals Evaluation of in vivo and in vitro Biological Activity of a Vibrio Cholerae 01 Hemolysin

2007 ◽  
Vol 30 (6) ◽  
pp. 250 ◽  
Author(s):  
Jose Arellano Galindo ◽  
Maria Guadalupe Rodriquez Angeles ◽  
Norma Valazquez Guadarrama ◽  
Enrique Santos Esteban ◽  
Silvia Giono Cerezo
Keyword(s):  
Vibrio Cholerae ◽  
Cell Cultures ◽  
Polyclonal Antibody ◽  
Elisa Test ◽  
Bacterial Lipopolysaccharide ◽  
Loop Model ◽  
Intestinal Epithelial ◽  
Ileal Loop

Purpose: To evaluate the hemolysin effect by ileal loop model produced by Vibrio cholerae O1 strains, compared with the cellular lysis or cytotoxic activity (CA) observed in cell culture. Method: We studied nine V. cholerae O1 strains, obtained during the Mexican outbreak of cholera (1990-1993), which had CA in Vero and CHO cells. Hemolysin was monitored with the hemolysis test. Titers of CA were calculated by CD50, and the association between CA and cholera toxin (CT) production was discarded by means of neutralization tests using an anti-CT polyclonal antibody. The CT production was measured with ELISA test. The LAL assay was performed in order to study relationships between the CA and bacterial lipopolysaccharide. Strains with CA were evaluated in rabbit and rat ileal loop models; hemorrhagic fluid was also measured. Tissues from ileal loop were included in paraffin to detect intestinal epithelial damage. Results: The hemolysin CA was not neutralized with the anti-CT polyclonal antibody. However, the associated factor of CA was heat labile. CA in cell cultures was not related to the bacterial lipopolysaccharide. The ileal loop test exhibited the presence of hemorrhagic tissue with inflammation. Conclusion: The V. cholerae O1 strains isolated were able to secrete hemolysin which, in turn, caused CA in cell cultures and produced the hemorrhagic and inflammatory effects observed in the ileal loop of rabbit and rat models.

scholarly journals Production of Monoclonal Antibodies Specific for the i and 1,2 Flagellar Antigens of Salmonella typhimurium and Characterization of Their Respective Epitopes

1998 ◽  
Vol 64 (12) ◽  
pp. 5033-5038 ◽  
Author(s):  
N. de Vries ◽  
K. A. Zwaagstra ◽  
J. H. J. Huis in’t Veld ◽  
F. van Knapen ◽  
F. G. van Zijderveld ◽  
...  
Keyword(s):  
Amino Acids ◽  
Monoclonal Antibodies ◽  
Salmonella Typhimurium ◽  
Immunosorbent Assay ◽  
Specific Antibody ◽  
Polyclonal Antibodies ◽  
Enzyme Linked Immunosorbent Assay ◽  
Specific Antibodies ◽  
Minimal Area

ABSTRACT Salmonella typhimurium expresses two antigenically distinct flagellins, each containing a different H antigen (i and 1,2), the combination of which is highly specific for this serotype. In this study, overlapping recombinant flagellin fragments were constructed from the fliC(H:i) and fljB (H:1,2) flagellin genes, and the expression products were tested for binding to H antigen-specific monoclonal and polyclonal antibodies. A minimal area, 86 amino acids for H:i and 102 amino acids for H:1,2, located in the central variable domain of each flagellin was required for the binding of serotype-specific antibodies, providing further evidence for the presence of a discontinuous H epitope. Two peptides comprising these areas were shown to be highly suitable for application as antigens in an enzyme-linked immunosorbent assay detecting S. typhimurium-specific antibody.

Unreliability of Current Serum Fibrin Degradation Product (FDP) Assays

1985 ◽  
Vol 53 (03) ◽  
pp. 301-302 ◽  
Author(s):  
P J Gaffney ◽  
M J Perry
Keyword(s):  
Monoclonal Antibodies ◽  
Serum Level ◽  
Degradation Product ◽  
Degradation Products ◽  
Polyclonal Antibodies ◽  
Fibrin Degradation Product ◽  
Serum Samples ◽  
Fibrinogen Degradation Products ◽  
A Value

SummaryPreviously, assays of fibrin-fibrinogen degradation products (FDP) had to be performed on serum samples. However, monoclonal antibodies (Mabs) are now available which permit the measurement of FDP directly in plasma. We have employed two Mabs, one monospecific for FDP originating from crosslinked fibrin and another panspecific for the FDP fraction, to determine normal FDP levels in plasma and serum. The monospecific Mab gave a value of 40 ng FDP/ml in plasma and 10 ng/ml in serum, while the serum level of FDP recorded using the panspecific Mab was >1000 ng/ml, at all the concentrations of thrombin employed. Similarly, when a solution of purified fibrinogen was treated with thrombin, the concentration of FDP present in the clot supernatant was >1000 ng/ml when assayed using the panspecific Mab. Thus during serum preparation as much as 75% of the native FDP is incorporated into the clot while in excess of 1000 ng/ml of laboratory generated FDP, probably incompletely polymerized fibrin, is measured using panspecific antisera. These data indicate that current FDP assays using polyclonal antibodies are not a reliable reflection of the FDP level generated in vivo. The use of FDP-specific Mabs which do not react with fibrinogen is recommended for future FDP assays performed directly on plasma.

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