ResearchPad - antibody-response https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[When two are better than one: Modeling the mechanisms of antibody mixtures]]> https://www.researchpad.co/article/elastic_article_14641 With the rise of new antibody combinations in therapeutic regimens, it is important to understand how antibodies work together as well as individually. Here, we investigate the specific case of monoclonal antibodies targeting a cancer-causing receptor or the influenza virus and develop a statistical mechanical framework that predicts the effectiveness of a mixture of antibodies. The power of this model lies in its ability to make a large number of predictions based on a limited amount of data. For example, once 10 antibodies have been individually characterized and their epitopes have been mapped, our model can predict how any of the 210 = 1024 combinations will behave. This predictive power can aid therapeutic efforts by assessing which combinations of antibodies will elicit the most effective response.

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<![CDATA[Low response in eliciting neuraminidase inhibition activity of sera among recipients of a split, monovalent pandemic influenza vaccine during the 2009 pandemic]]> https://www.researchpad.co/article/elastic_article_14470 Antibodies against influenza virus neuraminidase (NA) protein prevent releasing of the virus from host cells and spreading of infection foci and are considered the ‘second line of defence’ against influenza. Haemagglutinin inhibition antibody-low responders (HI-LRs) are present among influenza split vaccine recipients. The NA inhibition (NAI) antibody response in vaccinees is worth exploring, especially those in the HI-LRs population. We collected pre- and post-vaccination sera from 61 recipients of an inactivated, monovalent, split vaccine against A/H1N1pdm09 and acute and convalescent sera from 49 unvaccinated patients naturally infected with the A/H1N1pdm09 virus during the 2009 influenza pandemic. All samples were subjected to haemagglutinin inhibition (HI), NAI and neutralisation assays. Most paired sera from naturally infected patients exhibited marked elevation in the NAI activity, and seroconversion rates (SCR) among HI-LRs and HI-responders (HI-Rs) were 60% and 87%, respectively; however, those from vaccinees displayed low increase in the NAI activity, and the SCR among HI-LRs and HI-Rs were 0% and 12%, respectively. In both HI-LRs and HI-Rs, vaccination with the inactivated, monovalent, split vaccine failed to elicit the NAI activity efficiently in the sera of the naive population, compared with the natural infection. Hence, the improvement of influenza vaccines is warranted to elicit not only HI but also NAI antibodies.

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<![CDATA[Clinical role, optimal timing and frequency of serum infliximab and anti-infliximab antibody level measurements in patients with inflammatory bowel disease]]> https://www.researchpad.co/article/5989db50ab0ee8fa60bdc1f4

Background

Serum infliximab (IFX) and antibody-to-infliximab (ATI) levels are objective parameters, that may have a great role in the therapeutic decisions during maintenance biological therapy.

Research design and methods

48 inflammatory bowel disease patients receiving maintenance IFX therapy were prospectively enrolled and divided into adequate (complete remission N = 20) and inadequate responder (partial response, loss of response, dose escalation; N = 28) groups. Blood samples were collected just before (trough level, TL) and two (W2aTL) and six weeks (W6aTL) after the administration of IFX.

Results

Single measurement of ATI titer was insufficient for predicting therapeutic response due to transient expression of ATI, however, using the three points’ measurements, significant difference has been detected between the adequate and inadequate responder group (5.0% vs 35.7%; p = 0.016). The mean value of TL was significantly higher in the adequate responder group (3.11±1.64 vs.1.19±1.11; p<0.001) without further difference on the second and sixth week. Sensitivity and specificity for predicting the therapeutic response were 85.0% and 71.4% based on the cut-off value of TL 2.0 μg/ml.

Conclusion

Simultaneous measurement of serum IFX level prior to administration of regular IFX infusion and ATI titers significantly increase the diagnostic accuracy for the therapeutic decision in patients uncertainly responding to the therapy. The measurement of W2aTL and W6aTL levels did not result in further improvement in the prediction of therapeutic response.

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<![CDATA[Boosting subdominant neutralizing antibody responses with a computationally designed epitope-focused immunogen]]> https://www.researchpad.co/article/5c784fe8d5eed0c4840078de

Throughout the last several decades, vaccination has been key to prevent and eradicate infectious diseases. However, many pathogens (e.g., respiratory syncytial virus [RSV], influenza, dengue, and others) have resisted vaccine development efforts, largely because of the failure to induce potent antibody responses targeting conserved epitopes. Deep profiling of human B cells often reveals potent neutralizing antibodies that emerge from natural infection, but these specificities are generally subdominant (i.e., are present in low titers). A major challenge for next-generation vaccines is to overcome established immunodominance hierarchies and focus antibody responses on crucial neutralization epitopes. Here, we show that a computationally designed epitope-focused immunogen presenting a single RSV neutralization epitope elicits superior epitope-specific responses compared to the viral fusion protein. In addition, the epitope-focused immunogen efficiently boosts antibodies targeting the palivizumab epitope, resulting in enhanced neutralization. Overall, we show that epitope-focused immunogens can boost subdominant neutralizing antibody responses in vivo and reshape established antibody hierarchies.

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<![CDATA[Antibody responses to Plasmodium vivax Duffy binding and Erythrocyte binding proteins predict risk of infection and are associated with protection from clinical Malaria]]> https://www.researchpad.co/article/5c706793d5eed0c4847c7266

Background

The Plasmodium vivax Duffy Binding Protein (PvDBP) is a key target of naturally acquired immunity. However, region II of PvDBP, which contains the receptor-binding site, is highly polymorphic. The natural acquisition of antibodies to different variants of PvDBP region II (PvDBPII), including the AH, O, P and Sal1 alleles, the central region III-V (PvDBPIII-V), and P. vivax Erythrocyte Binding Protein region II (PvEBPII) and their associations with risk of clinical P. vivax malaria are not well understood.

Methodology

Total IgG and IgG subclasses 1, 2, and 3 that recognize four alleles of PvDBPII (AH, O, P, and Sal1), PvDBPIII-V and PvEBPII were measured in samples collected from a cohort of 1 to 3 year old Papua New Guinean (PNG) children living in a highly endemic area of PNG. The levels of binding inhibitory antibodies (BIAbs) to PvDBPII (AH, O, and Sal1) were also tested in a subset of children. The association of presence of IgG with age, cumulative exposure (measured as the product of age and malaria infections during follow-up) and prospective risk of clinical malaria were evaluated.

Results

The increase in antigen-specific total IgG, IgG1, and IgG3 with age and cumulative exposure was only observed for PvDBPII AH and PvEBPII. High levels of total IgG and predominant subclass IgG3 specific for PvDBPII AH were associated with decreased incidence of clinical P. vivax episodes (aIRR = 0.56–0.68, P≤0.001–0.021). High levels of total IgG and IgG1 to PvEBPII correlated strongly with protection against clinical vivax malaria compared with IgGs against all PvDBPII variants (aIRR = 0.38, P<0.001). Antibodies to PvDBPII AH and PvEBPII showed evidence of an additive effect, with a joint protective association of 70%.

Conclusion

Antibodies to the key parasite invasion ligands PvDBPII and PvEBPII are good correlates of protection against P. vivax malaria in PNG. This further strengthens the rationale for inclusion of PvDBPII in a recombinant subunit vaccine for P. vivax malaria and highlights the need for further functional studies to determine the potential of PvEBPII as a component of a subunit vaccine for P. vivax malaria.

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<![CDATA[Late effects of total body irradiation on hematopoietic recovery and immune function in rhesus macaques]]> https://www.researchpad.co/article/5c6dc9f6d5eed0c48452a5fd

While exposure to radiation can be lifesaving in certain settings, it can also potentially result in long-lasting adverse effects, particularly to hematopoietic and immune cells. This study investigated hematopoietic recovery and immune function in rhesus macaques Cross-sectionally (at a single time point) 2 to 5 years after exposure to a single large dose (6.5 to 8.4 Gray) of total body radiation (TBI) derived from linear accelerator-derived photons (2 MeV, 80 cGy/minute) or Cobalt 60-derived gamma irradiation (60 cGy/min). Hematopoietic recovery was assessed through measurement of complete blood counts, lymphocyte subpopulation analysis, and thymus function assessment. Capacity to mount specific antibody responses against rabies, Streptococcus pneumoniae, and tetanus antigens was determined 2 years after TBI. Irradiated macaques showed increased white blood cells, decreased platelets, and decreased frequencies of peripheral blood T cells. Effects of prior radiation on production and export of new T cells by the thymus was dependent on age at the time of analysis, with evidence of interaction with radiation dose for CD8+ T cells. Irradiated and control animals mounted similar mean antibody responses to proteins from tetanus and rabies and to 10 of 11 serotype-specific pneumococcal polysaccharides. However, irradiated animals uniformly failed to make antibodies against polysaccharides from serotype 5 pneumococci, in contrast to the robust responses of non-irradiated controls. Trends toward decreased serum levels of anti-tetanus IgM and slower peak antibody responses to rabies were also observed. Taken together, these data show that dose-related changes in peripheral blood cells and immune responses to both novel and recall antigens can be detected 2 to 5 years after exposure to whole body radiation. Longer term follow-up data on this cohort and independent validation will be helpful to determine whether these changes persist or whether additional changes become evident with increasing time since radiation, particularly as animals begin to develop aging-related changes in immune function.

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<![CDATA[Bivalent oral cholera vaccination induces a memory B cell response to the V. cholerae O1-polysaccharide antigen in Haitian adults]]> https://www.researchpad.co/article/5c5ca2c6d5eed0c48441eaae

The bivalent killed whole-cell oral cholera vaccine (BivWC) is being increasingly used to prevent cholera. The presence of O-antigen-specific memory B cells (MBC) has been associated with protective immunity against cholera, yet MBC responses have not been evaluated after BivWC vaccination. To address this knowledge gap, we measured V. cholerae O1-antigen MBC responses following BivWC vaccination. Adults in St. Marc, Haiti, received 2 doses of the BivWC vaccine, Shanchol, two weeks apart. Participants were invited to return at days 7, 21, 44, 90, 180 and 360 after the initial vaccination. Serum antibody and MBC responses were assessed at each time-point before and following vaccination. We observed that vaccination with BivWC resulted in significant O-antigen specific MBC responses to both Ogawa and Inaba serotypes that were detected by day 21 and remained significantly elevated over baseline for up to 12 months following vaccination. The BivWC oral cholera vaccine induces durable MBC responses to the V. cholerae O1-antigen. This suggests that long-term protection observed following vaccination with BivWC could be mediated or maintained by MBC responses.

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<![CDATA[Community-level chlamydial serology for assessing trachoma elimination in trachoma-endemic Niger]]> https://www.researchpad.co/article/5c58d649d5eed0c484031a9e

Background

Program decision-making for trachoma elimination currently relies on conjunctival clinical signs. Antibody tests may provide additional information on the epidemiology of trachoma, particularly in regions where it is disappearing or elimination targets have been met.

Methods

A cluster-randomized trial of mass azithromycin distribution strategies for trachoma elimination was conducted over three years in a mesoendemic region of Niger. Dried blood spots were collected from a random sample of children aged 1–5 years in each of 24 study communities at 36 months after initiation of the intervention. A multiplex bead assay was used to test for antibodies to two Chlamydia trachomatis antigens, Pgp3 and CT694. We compared seropositivity to either antigen to clinical signs of active trachoma (trachomatous inflammation—follicular [TF] and trachomatous inflammation—intense [TI]) at the individual and cluster level, and to ocular chlamydia prevalence at the community level.

Results

Of 988 children with antibody data, TF prevalence was 7.8% (95% CI 6.1 to 9.5) and TI prevalence was 1.6% (95% CI 0.9 to 2.6). The overall prevalence of antibody positivity to Pgp3 was 27.2% (95% CI 24.5 to 30), and to CT694 was 23.7% (95% CI 21 to 26.2). Ocular chlamydia infection prevalence was 5.2% (95% CI 2.8 to 7.6). Seropositivity to Pgp3 and/or CT694 was significantly associated with TF at the individual and community level and with ocular chlamydia infection and TI at the community level. Older children were more likely to be seropositive than younger children.

Conclusion

Seropositivity to Pgp3 and CT694 correlates with clinical signs and ocular chlamydia infection in a mesoendemic region of Niger.

Trial registration

ClinicalTrials.gov NCT00792922.

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<![CDATA[Environmental complexity: A buffer against stress in the domestic chick]]> https://www.researchpad.co/article/5c466559d5eed0c484518bb5

Birds kept in commercial production systems can be exposed to multiple stressors from early life and this alters the development of different morphological, immunological and behavioural indicators. We explore the hypothesis that provision of a complex environment during early life, better prepares birds to cope with stressful events as well as buffers them against future unpredictable stressful episodes. In this study, 96 one day old pullets were randomly distributed in eight pens (12 birds/pen). Half of the chicks (N = 48) were assigned to a Complex Environment (CENV: with perches, a dark brooder etc.) the others to a Simple Environment (SENV: without enrichment features). Half of the birds from each of these treatments were assigned to a No Stress (NSTR, 33°C) or to an acute Cold Stress (CSTR, 18–20°C) treatment during six hours on their second day of life. At four weeks of age, chicks with these four different backgrounds were exposed to an Intermittent Stressful Challenges Protocol (ISCP). In an immunological test indicative of pro-inflammatory status Phytohemagglutinin-P (PHA-P), the response of CSTR birds was ameliorated by rearing chicks in a CENV as they had a similar response to NSTR chicks and a significantly better pro-inflammatory response than those CSTR birds reared in a SENV (five days after the CSTR treatment was applied). A similar better response when coping with new challenges (the ISCP) was observed in birds reared in a CENV compared to those from a SENV. Birds reared in the CENV had a lower heterophil/lymphocyte ratio after the ISCP than birds reared in SENV, independently of whether or not they had been exposed to CSTR early in life. No effects of stress on general behaviour were detected, however, the provision of a CENV increased resting behaviour, which may have favoured stress recover. Additionally, we found that exposure to cold stress at an early age might have rendered birds more vulnerable to future stressful events. CSTR birds had lower humoral immune responses (sheep red blood cells induced antibodies) after the ISCP and started using elevated structures in the CENV later compared to their NSTR conspecifics. Our study reflects the importance of the early provision of a CENV in commercial conditions to reduce negative stress-related effects. Within the context of the theory of adaptive plasticity, our results suggest that the early experience of the birds had long lasting effects on the modulation of their phenotypes.

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<![CDATA[A plant-derived VLP influenza vaccine elicits a balanced immune response even in very old mice with co-morbidities]]> https://www.researchpad.co/article/5c40f7a9d5eed0c484386566

Background

The elderly are at high risk from influenza, in part because immunity wanes with age and through the accumulation of comorbidities. A novel plant-derived virus-like-particle (VLP) vaccine bearing influenza hemagglutinin can induce a balanced humoral and cellular response in old mice (16–18 months) while split virion vaccines elicit mostly antibodies. Because mice also collect comorbidities and lose immune competence as they age, we wished to determine how the plant-derived VLP vaccine would perform in animals approaching the end of their life-span.

Materials and methods

Old (24–26 months) female BALB/c mice received two intramuscular doses of H1-VLP vaccine, an inactivated H1N1 vaccine (IIV) (both based on A/H1N1/California/07/09) (3μg each) or PBS. Serum was collected on day 42 and humoral responses were measured by enzyme-linked immunosorbent assay (ELISA), microneutralization (MN) and hemagglutination inhibition (HI) assays. Influenza-specific splenocyte CD4+ & CD8+ T cell responses were measured by flow cytometry. Full body computed tomography (CT) and structured necropsies were performed on day 42. Comorbidities including reduced lung volume (kyphosis), masses, abscesses, etc. were assessed using a standard scoring system (1–21) and mice with scores ≥5 were considered to have important comorbidities.

Results

Overall, 53.3% of the animals had significant comorbidities. Three weeks post-boost, HI and MN titres were mostly undetectable but ELISA titres were significantly higher in the H1-VLP animals compared to the IIV group (GMT (95% CI): 961 (427, 2163) vs 425 (200, 903): p = 0.03). Both CD4+(TNFα, IFNγ) and CD8+ (IFNγ) T cell responses were also greater in the H1-VLP group than the IIV.

Conclusions

Even in very old mice with comorbidities, the plant-made H1-VLP vaccine elicited a stronger and more balanced immune response than IIV. Animals with fewer comorbidities tended to have the better composite (humoral and cellular) responses. These novel vaccines have the potential to address some of the limitations of current vaccines in the elderly.

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<![CDATA[The effects of prednisolone treatment on serological responses and lipid profiles in Ethiopian leprosy patients with Erythema Nodosum Leprosum reactions]]> https://www.researchpad.co/article/5c2fcf50d5eed0c484a6e0c2

Background

Erythema nodosum leprosum (ENL) is a systemic inflammatory complication occurring mainly in patients with lepromatous leprosy (LL) and borderline lepromatous leprosy (BL). Prednisolone is widely used for treatment of ENL reactions. However, it has been reported that prolonged treatment with prednisolone increases the risk for prednisolone-induced complications such as osteoporosis, diabetes, cataract and arteriosclerosis. It has been speculated that perhaps these complications result from lipid profile alterations by prednisolone. The effects of extended prednisolone treatment on lipid profiles in ENL patients have not been studied in leprosy patients with ENL reactions. Therefore, in this study we conducted a case-control study to investigate the changes in lipid profiles and serological responses in Ethiopian patients with ENL reaction after prednisolone treatment.

Methods

A prospective matched case–control study was employed to recruit 30 patients with ENL and 30 non-reactional LL patient controls at ALERT Hospital, Ethiopia. Blood samples were obtained from each patient with ENL reaction before and after prednisolone treatment as well as from LL controls. The serological host responses to PGL-1, LAM and Ag85 M. leprae antigens were measured by ELISA. Total cholesterol (TC), triglyceride (TG), high density lipoprotein (HDL) and low density lipoprotein (LDL) were measured by spectrophotometric method.

Results

The host antibody response to M. leprae PGL-1, LAM and Ag85 antigens were significantly reduced in patients with ENL reactions compared to LL controls after treatment. Comparison between patients with acute and chronic ENL showed that host-response to PGL-1 was significantly reduced in chronic ENL after prednisolone treatment. Untreated patients with ENL reactions had low lipid concentration compared to LL controls. However, after treatment, both groups had comparable lipid profiles except for LDL, which was significantly higher in patients with ENL reaction. Comparison within the ENL group before and after treatment showed that prednisolone significantly increased LDL and HDL levels in ENL patients and this was more prominent in chronic ENL than in acute patients with ENL.

Conclusion

The significantly increased prednisolone-induced LDL and TG levels, particularly in patients with chronic ENL reactions, is a concern in the use of prednisolone for extended periods in ENL patients. The findings highlight the importance of monitoring lipid profiles during treatment of patients to minimize the long-term risk of prednisolone-induced complications.

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<![CDATA[IgG1 and IgG4 antibodies against Aedes aegypti salivary proteins and risk for dengue infections]]> https://www.researchpad.co/article/5c36679fd5eed0c4841a5e0d

Dengue virus (DENV) is an arbovirus responsible for a significant number of deaths in Latin America. This virus is transmitted through the bite of Aedes aegypti, the main mosquito vector, and Ae. albopictus. During blood uptake, the mosquito injects its saliva into the host to facilitate the feeding process. Mosquito saliva contains potent immunogens capable of inducing antibody production directly related to mosquito bite exposure intensity and disease risk. In this study, we first determined the DENV infection status by two different DENV non-structural protein 1 (NS1) based rapid tests and qRT-PCR, then measured the levels of IgG1 and IgG4 antibodies against salivary proteins of Ae. aegypti female mosquitoes in volunteers living in a dengue endemic area. Our results show that people with a positive DENV diagnosis present higher levels of IgG4 antibodies than people with a negative diagnostic test, and that these antibody levels were higher in people with secondary DENV infections. With this study, we show that detection of IgG4 antibodies against mosquito saliva may be a reliable method to evaluate the risk of dengue infection.

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<![CDATA[Vectored delivery of anti-SIV envelope targeting mAb via AAV8 protects rhesus macaques from repeated limiting dose intrarectal swarm SIVsmE660 challenge]]> https://www.researchpad.co/article/5c117b9ed5eed0c484699e2f

Gene based delivery of immunoglobulins promises to safely and durably provide protective immunity to individuals at risk of acquiring infectious diseases such as HIV. We used a rhesus macaque animal model to optimize delivery of naturally-arising, autologous anti-SIV neutralizing antibodies expressed by Adeno-Associated Virus 8 (AAV8) vectors. Vectored transgene expression was confirmed by quantitation of target antibody abundance in serum and mucosal surfaces. We tested the expression achieved at varying doses and numbers of injections. Expression of the transgene reached a saturation at about 2 x 1012 AAV8 genome copies (gc) per needle-injection, a physical limitation that may not scale clinically into human trials. In contrast, expression increased proportionately with the number of injections. In terms of anti-drug immunity, anti-vector antibody responses were universally strong, while those directed against the natural transgene mAb were detected in only 20% of animals. An anti-transgene antibody response was invariably associated with loss of detectable plasma expression of the antibody. Despite having atypical glycosylation profiles, transgenes derived from AAV-directed muscle cell expression retained full functional activity, including mucosal accumulation, in vitro neutralization, and protection against repeated limiting dose SIVsmE660 swarm challenge. Our findings demonstrate feasibility of a gene therapy-based passive immunization strategy against infectious disease, and illustrate the potential for the nonhuman primate model to inform clinical AAV-based approaches to passive immunization.

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<![CDATA[Antibody responses to Zika virus proteins in pregnant and non-pregnant macaques]]> https://www.researchpad.co/article/5c06f04ad5eed0c484c6d62b

The specificity of the antibody response against Zika virus (ZIKV) is not well-characterized. This is due, in part, to the antigenic similarity between ZIKV and closely related dengue virus (DENV) serotypes. Since these and other similar viruses co-circulate, are spread by the same mosquito species, and can cause similar acute clinical syndromes, it is difficult to disentangle ZIKV-specific antibody responses from responses to closely-related arboviruses in humans. Here we use high-density peptide microarrays to profile anti-ZIKV antibody reactivity in pregnant and non-pregnant macaque monkeys with known exposure histories and compare these results to reactivity following DENV infection. We also compare cross-reactive binding of ZIKV-immune sera to the full proteomes of 28 arboviruses. We independently confirm a purported ZIKV-specific IgG antibody response targeting ZIKV nonstructural protein 2B (NS2B) that was recently reported in ZIKV-infected people and we show that antibody reactivity in pregnant animals can be detected as late as 127 days post-infection (dpi). However, we also show that these responses wane over time, sometimes rapidly, and in one case the response was elicited following DENV infection in a previously ZIKV-exposed animal. These results suggest epidemiologic studies assessing seroprevalence of ZIKV immunity using linear epitope-based strategies will remain challenging to interpret due to susceptibility to false positive results. However, the method used here demonstrates the potential for rapid profiling of proteome-wide antibody responses to a myriad of neglected diseases simultaneously and may be especially useful for distinguishing antibody reactivity among closely related pathogens.

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<![CDATA[The adjuvant AlhydroGel elicits higher antibody titres than AddaVax when combined with HIV-1 subtype C gp140 from CAP256]]> https://www.researchpad.co/article/5c215190d5eed0c4843faca8

With the HIV-1 epidemic in southern Africa still rising, a prophylactic vaccine against the region’s most prolific subtype (subtype C) would be a significant step forward. In this paper we report on the effect of 2 different adjuvants, AddaVax and AlhydroGel, formulated with HIV-1 subtype C gp140, on the development of binding and neutralising antibody titres in rabbits. AddaVax is a squalene-based oil-in-water nano-emulsion (similar to MF59) which can enhance both cellular and humoral immune responses, whilst AlhydroGel (aluminium hydroxide gel) mainly drives a Th2 response. The gp140 gene tested was derived from the superinfecting virus (SU) from participant CAP256 in the CAPRISA 002 Acute infection cohort. The furin cleavage site of the Env protein was replaced with a flexible linker and an I559P mutation introduced. Lectin affinity purified soluble Env protein was mainly trimeric as judged by molecular weight using BN-PAGE and contained intact broadly neutralising epitopes for the V3-glycan supersite (monoclonal antibodies PGT128 and PGT135), the CD4 binding site (VRC01) and the V2-glycan (PG9) but not for the trimer-specific monoclonal antibodies PG16, PGT145 and CAP256-VRC26_08. When this soluble Env protein was tested in rabbits, AlhydroGel significantly enhanced soluble Env and V1V2 binding antibodies when compared to AddaVax. Finally, AlhydroGel resulted in significantly higher neutralization titres for a subtype C Tier 1A virus (MW965.26) and increased neutralization breadth to Tier 1A and 1B viruses. However, no autologous Tier 2 neutralisation was observed. These data suggest that adjuvant selection is critical for developing a successful vaccine and AlhydroGel should be further investigated. Additional purification of trimeric native-like CAP256 Env and/or priming with DNA or MVA might enhance the induction of neutralizing antibodies and possible Tier 2 HIV-1 neutralisation.

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<![CDATA[Human antibody reaction against recombinant salivary proteins of Phlebotomus orientalis in Eastern Africa]]> https://www.researchpad.co/article/5c1028b4d5eed0c484247e0e

Background

Phlebotomus orientalis is a vector of Leishmania donovani, the causative agent of life threatening visceral leishmaniasis spread in Eastern Africa. During blood-feeding, sand fly females salivate into the skin of the host. Sand fly saliva contains a large variety of proteins, some of which elicit specific antibody responses in the bitten hosts. To evaluate the exposure to sand fly bites in human populations from disease endemic areas, we tested the antibody reactions of volunteers' sera against recombinant P. orientalis salivary antigens.

Methodology/Principal findings

Recombinant proteins derived from sequence data on P. orientalis secreted salivary proteins, were produced using either bacterial (five proteins) or mammalian (four proteins) expression systems and tested as antigens applicable for detection of anti-P. orientalis IgG in human sera. Using these recombinant proteins, human sera from Sudan and Ethiopia, countries endemic for visceral leishmaniasis, were screened by ELISA and immunoblotting to identify the potential markers of exposure to P. orientalis bites. Two recombinant proteins; mAG5 and mYEL1, were identified as the most promising antigens showing high correlation coefficients as well as good specificity in comparison to the whole sand fly salivary gland homogenate. Combination of both proteins led to a further increase of correlation coefficients as well as both positive and negative predictive values of P. orientalis exposure.

Conclusions/Significance

This is the first report of screening human sera for anti-P. orientalis antibodies using recombinant salivary proteins. The recombinant salivary proteins mYEL1 and mAG5 proved to be valid antigens for screening human sera from both Sudan and Ethiopia for exposure to P. orientalis bites. The utilization of equal amounts of these two proteins significantly increased the capability to detect anti-P. orientalis antibody responses.

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<![CDATA[Nucleocapsid protein-based vaccine provides protection in mice against lethal Crimean-Congo hemorrhagic fever virus challenge]]> https://www.researchpad.co/article/5b60074b463d7e39c55261ff

Crimean-Congo hemorrhagic fever (CCHF) is an acute, often fatal viral disease characterized by rapid onset of febrile symptoms followed by hemorrhagic manifestations. The etiologic agent, CCHF orthonairovirus (CCHFV), can infect several mammals in nature but only seems to cause clinical disease in humans. Over the past two decades there has been an increase in total number of CCHF case reports, including imported CCHF patients, and an expansion of CCHF endemic areas. Despite its increased public health burden there are currently no licensed vaccines or treatments to prevent CCHF. We here report the development and assessment of the protective efficacy of an adenovirus (Ad)-based vaccine expressing the nucleocapsid protein (N) of CCHFV (Ad-N) in a lethal immunocompromised mouse model of CCHF. The results show that Ad-N can protect mice from CCHF mortality and that this platform should be considered for future CCHFV vaccine strategies.

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<![CDATA[Immunogenicity and efficacy following sequential parenterally-administered doses of Salmonella Enteritidis COPS:FliC glycoconjugates in infant and adult mice]]> https://www.researchpad.co/article/5b28ae90463d7e0f49b27045

In sub-Saharan Africa, invasive nontyphoidal Salmonella (iNTS) infections with serovars S. Enteritidis, S. Typhimurium and I 4,[5],12:i:- are widespread in children < 5 years old. Development of an efficacious vaccine would provide an important public health tool to prevent iNTS disease in this population. Glycoconjugates of S. Enteritidis core and O-polysaccharide (COPS) coupled to the homologous serovar phase 1 flagellin protein (FliC) were previously shown to be immunogenic and protected adult mice against death following challenge with a virulent Malian S. Enteritidis blood isolate. This study extends these observations to immunization of mice in early life and also assesses protection with partial and full regimens. Anti-COPS and anti-FliC serum IgG titers were assessed in infant and adult mice after immunization with 1, 2 or 3 doses of S. Enteritidis COPS:FliC alone or co-formulated with aluminum hydroxide or monophosphoryl lipid A (MPL) adjuvants. S. Enteritidis COPS:FliC was immunogenic in both age groups, although the immune responses were quantitatively lower in infants. Kinetics of antibody production were similar for the native and adjuvanted formulations after three doses; conjugates formulated with MPL elicited significantly increased anti-COPS IgG titers in adult but not infant mice. Nevertheless, robust protection against S. Enteritidis challenge was seen for all three formulations when three doses were given either during infancy or as adults. We further found that significant protection could be achieved with two COPS:FliC doses, despite elicitation of modest serum anti-COPS IgG antibody titers. These findings guide potential immunization strategies that may be translated to develop a human pediatric iNTS vaccine for sub-Saharan Africa.

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<![CDATA[Analysis of the effect of promoter type and skin pretreatment on antigen expression and antibody response after gene gun-based immunization]]> https://www.researchpad.co/article/5b28b3fb463d7e129299938c

Monoclonal antibodies (mAbs) have enabled numerous basic research discoveries and therapeutic approaches for many protein classes. However, there still exist a number of target classes, such as multi-pass membrane proteins, for which antibody discovery is difficult, due in part to lack of high quality, recombinant protein. Here we describe the impact of several parameters on antigen expression and the development of mAbs against human claudin 4 (CLDN4), a potential multi-indication cancer target. Using gene gun-based DNA delivery and bioluminescence imaging, we optimize promoter type by comparing expression profiles of four robust in vivo promoters. In addition, we observe that most vectors rapidly lose expression, ultimately reaching almost background levels by three days post-delivery. Recognizing this limitation, we next explored skin pretreatment strategies as an orthogonal method to further boost the efficiency of mAb generation. We show that SDS pretreatment can boost antigen expression, but fails to significantly increase mAb discovery efficiency. In contrast, we find that sandpaper pretreatment yields 5-fold more FACS+ anti-CLDN4 hybridomas, without impacting antigen expression. Our findings coupled with other strategies to improve DNA immunizations should improve the success of mAb discovery against other challenging targets and enable the generation of critical research tools and therapeutic candidates.

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<![CDATA[IgG1 as a Potential Biomarker of Post-chemotherapeutic Relapse in Visceral Leishmaniasis, and Adaptation to a Rapid Diagnostic Test]]> https://www.researchpad.co/article/5989daecab0ee8fa60bbfa51

Background

Visceral leishmaniasis (VL), caused by protozoa of the Leishmania donovani complex, is a widespread parasitic disease of great public health importance; without effective chemotherapy symptomatic VL is usually fatal. Distinction of asymptomatic carriage from progressive disease and the prediction of relapse following treatment are hampered by the lack of prognostic biomarkers for use at point of care.

Methodology/Principal Findings

All IgG subclass and IgG isotype antibody levels were determined using unpaired serum samples from Indian and Sudanese patients with differing clinical status of VL, which included pre-treatment active VL, post-treatment cured, post-treatment relapsed, and post kala-azar dermal leishmaniasis (PKDL), as well as seropositive (DAT and/or rK39) endemic healthy controls (EHCs) and seronegative EHCs. L. donovani antigen-specific IgG1 levels were significantly elevated in relapsed versus cured VL patients (p<0.0001). Using paired Indian VL sera, consistent with the known IgG1 half-life, IgG1 levels had not decreased significantly at day 30 after the start of treatment (p = 0.8304), but were dramatically decreased by 6 months compared to day 0 (p = 0.0032) or day 15 (p<0.0001) after start of treatment. Similarly, Sudanese sera taken soon after treatment did not show a significant change in the IgG1 levels (p = 0.3939). Two prototype lateral flow immunochromatographic rapid diagnostic tests (RDTs) were developed to detect IgG1 levels following VL treatment: more than 80% of the relapsed VL patients were IgG1 positive; at least 80% of the cured VL patients were IgG1 negative (p<0.0001).

Conclusions/Significance

Six months after treatment of active VL, elevated levels of specific IgG1 were associated with treatment failure and relapse, whereas no IgG1 or low levels were detected in cured VL patients. A lateral flow RDT was successfully developed to detect anti-Leishmania IgG1 as a potential biomarker of post-chemotherapeutic relapse.

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