共查询到20条相似文献,搜索用时 15 毫秒
1.
Rosales-Mendoza S Soria-Guerra RE López-Revilla R Moreno-Fierros L Alpuche-Solís AG 《Plant cell reports》2008,27(1):79-84
Diarrheal diseases caused by Vibrio cholerae and enterotoxigenic Escherichia coli (ETEC) are worldwide health problems that might be prevented with vaccines based on edible plants expressing the B subunit
from either the cholera toxin (CTB) or the E. coli heat labile toxin (LTB). In this work we analyzed the immunity induced in Balb/c mice by ingestion of three weekly doses
of 10 μg of LTB derived from transgenic carrot material. Although the anti-LTB serum immunoglobulin G (IgG) and intestinal
IgA antibody responses were higher with 10 μg-doses of pure bacterial recombinant LTB (rLTB), the transgenic carrot material
also elicited significant serum and intestinal antibody responses. Serum anti-LTB IgG1 antibodies predominated over IgG2a
antibodies, suggesting that mainly Th2 responses were induced. A decrease of intestinal fluid accumulation after cholera toxin
challenge was observed in mice immunized with either rLTB or LTB-containing carrot material. These results demonstrate that
ingestion of carrot-derived LTB induces antitoxin systemic and intestinal immunity in mice and suggest that transgenic carrots
expressing LTB may be used as an effective edible vaccine against cholera and ETEC diarrhea in humans. 相似文献
2.
Rosales-Mendoza S Alpuche-Solís AG Soria-Guerra RE Moreno-Fierros L Martínez-González L Herrera-Díaz A Korban SS 《The Plant journal : for cell and molecular biology》2009,57(1):45-54
Enterotoxigenic Escherichia coli (ETEC) strains are important pathogens in developing countries. Some vaccine formulations containing the heat labile toxin B subunit (LTB) have been used in clinical trials; however, the induction of neutralizing antibodies against the heat-stable toxin (ST), a poor immunogenic peptide, is necessary, as most ETEC strains can produce both toxins. In this study, a plant optimized synthetic gene encoding for the LTB-ST fusion protein has been introduced into plastids of tobacco leaf tissues, using biolistic microprojectile bombardment, in an effort to develop a single plant-based candidate vaccine against both toxins. Transplastomic tobacco plants carrying the LTB-ST transgene have been recovered. Transgene insertion into the plastid was confirmed by both PCR and Southern blot analysis. GM1-ELISA revealed that the LTB-ST fusion protein retained its oligomeric structure, and displayed antigenic determinants for both LTB and ST. Western blot analysis, using LTB antisera, confirmed the presence of a 17-KDa protein in transplastomic lines, with the correct antigenicity of the fusion protein. Expression levels of this fusion protein in different lines reached up to 2.3% total soluble protein. Oral immunization of mice with freeze-dried transplastomic tobacco leaves led to the induction of both serum and mucosal LTB-ST specific antibodies. Following cholera toxin challenge, a decrease of intestinal fluid accumulation was observed in mice immunized with LTB-ST-containing tobacco. These findings suggest that tobacco plants expressing LTB-ST could serve as a plant-based candidate vaccine model providing broad-spectrum protection against ETEC-induced diarrhoeal disease. 相似文献
3.
Abdoreza Davoodi‐Semiromi Melissa Schreiber Samson Nalapalli Dheeraj Verma Nameirakpam D. Singh Robert K. Banks Debopam Chakrabarti Henry Daniell 《Plant biotechnology journal》2010,8(2):223-242
Cholera and malaria are major diseases causing high mortality. The only licensed cholera vaccine is expensive; immunity is lost in children within 3 years and adults are not fully protected. No vaccine is yet available for malaria. Therefore, in this study, the cholera toxin‐B subunit (CTB) of Vibrio cholerae fused to malarial vaccine antigens apical membrane antigen‐1 (AMA1) and merozoite surface protein‐1 (MSP1) was expressed in lettuce and tobacco chloroplasts. Southern blot analysis confirmed homoplasmy and stable integration of transgenes. CTB‐AMA1 and CTB‐MSP1 fusion proteins accumulated up to 13.17% and 10.11% (total soluble protein, TSP) in tobacco and up to 7.3% and 6.1% (TSP) in lettuce, respectively. Nine groups of mice (n = 10/group) were immunized subcutaneously (SQV) or orally (ORV) with purified antigens or transplastomic tobacco leaves. Significant levels of antigen‐specific antibody titres of immunized mice completely inhibited proliferation of the malarial parasite and cross‐reacted with the native parasite proteins in immunoblots and immunofluorescence studies. Protection against cholera toxin challenge in both ORV (100%) and SQV (89%) mice correlated with CTB‐specific titres of intestinal, serum IgA and IgG1 in ORV and only IgG1 in SQV mice, but no other immunoglobulin. Increasing numbers of interleukin‐10+ T cell but not Foxp3+ regulatory T cells, suppression of interferon‐γ and absence of interleukin‐17 were observed in protected mice, suggesting that immunity is conferred via the Tr1/Th2 immune response. Dual immunity against two major infectious diseases provided by chloroplast‐derived vaccine antigens for long‐term (>300 days, 50% of mouse life span) offers a realistic platform for low cost vaccines and insight into mucosal and systemic immunity. 相似文献
4.
A Chimeric protein of CFA/I,CS6 subunits and LTB/STa toxoid protects immunized mice against enterotoxigenic Escherichia coli 下载免费PDF全文
Narges Zeinalzadeh Ali Hatef Salmanian Goli Goujani Jafar Amani Ghasem Ahangari Asal Akhavian Mahyat Jafari 《Microbiology and immunology》2017,61(7):272-279
5.
You J Xu Y He M McAllister TA Thacker PA Li X Wang T Jin L 《Applied microbiology and biotechnology》2011,89(6):1885-1893
Currently available enterotoxigenic Escherichia coli (ETEC) vaccines are based on colonization factors and/or the heat-labile enterotoxin B subunit (LTB). However, the induction
of antitoxic responses against heat-stable enterotoxin a (STa) and b (STb) has merit as these two poorly immunogenic toxins
are frequently associated with ETEC strains. In this study, we genetically constructed a trivalent enterotoxin fusion protein
(STa–LTB–STb, abbreviated to SLS) in an effort to develop a single toxoid containing these three enterotoxins for vaccination
against ETEC. Mutagenesis at one disulfide-bridge-forming cysteine in STa led to a dramatic reduction in the STa toxicity
of SLS; however, the fusion peptide retained the STb-associated toxicity. Immunization of mice with SLS protein elicited significant
antibody responses to LTB, STa, and STb. Significantly, the mice antisera were able to neutralize the biological activity
of both STa and STb. In the experiment to assess the protective effect of SLS immunization, the mortality of mice receiving
SLS was significantly lower than their control cohorts (P < 0.01) after intraperitoneal challenge with ETEC. These results show that the trivalent fusion enterotoxin SLS has the potential
to serve as a useful toxin-based vaccine against ETEC-induced diarrheal disease via a single immunogen. 相似文献
6.
Enterotoxigenic Escherichia coli (ETEC) is an important cause of diarrheal disease and deaths among children in developing countries and the major cause of
traveler's diarrhea (TD). Since surface protein colonization factors (CFs) of ETEC are important for pathogenicity and immune
protection is mainly mediated by locally produced IgA antibodies in the gut, much effort has focused on the development of
an oral CF-based vaccine. The most extensively studied ETEC candidate vaccine is the rCTB-CF ETEC vaccine, containing recombinantly
produced cholera B subunit and the most commonly encountered ETEC CFs on the surface of whole inactivated bacteria. Initial
clinical trials with this vaccine showed significant immune responses against the key antigens in different age groups in
Bangladesh and Egypt and protection against more severe TD in Western travelers. However, when tested in a phase-III trial
in Egyptian infants, the protective efficacy of the vaccine was found to be low, indicating the need to improve the immunogenicity
of the vaccine, e.g., by increasing the levels of the protective antigens. This review describes different strategies for
the construction of recombinant nontoxigenic E. coli and Vibrio cholerae candidate vaccine strains over-expressing higher amounts of ETEC CFs than clinical ETEC isolates selected to produce high
levels of the respective CF, e.g., those ETEC strains which have been used in the rCTB-CF ETEC vaccine. Several different
expression vectors containing the genes responsible for the expression and assembly of the examined CFs, all downstream of
the powerful tac promoter, which could be maintained either with or without antibiotic selection, were constructed. Expression from the tac promoter was under the control of the lacI
q repressor present on the plasmids. Following induction with isopropyl-β-d-thiogalactopyranoside, candidate vaccine strains over-expressing single CFs, unnatural combinations of two CFs, and also
hybrid forms of ETEC CFs were produced. Specific monoclonal antibodies against the major subunits of the examined CF were
used to quantify the amount of the surface-expressed CF by a dot-blot assay and inhibition ELISA. Oral immunization with formalin-
or phenol-inactivated recombinant bacteria over-expressing the CFs was found to induce significantly higher antibody responses
compared to immunization with the previously used vaccine strains. We therefore conclude that our constructs may be useful
as candidate strains in an oral whole-cell inactivated CF ETEC vaccine. 相似文献
7.
Rosales-Mendoza S Soria-Guerra RE Moreno-Fierros L Govea-Alonso DO Herrera-Díaz A Korban SS Alpuche-Solís ÁG 《Plant cell reports》2011,30(6):1145-1152
Enterotoxigenic Escherichia coli (ETEC) is one of the main causative agents of diarrhea in infants and for travelers. Inclusion of a heat-stable (ST) toxin into vaccine formulations is mandatory as most ETEC strains can produce both heat-labile (LT) and ST enterotoxins. In this study, a genetic fusion gene encoding for an LTB:ST protein has been constructed and transferred into tobacco via Agrobacterium tumefaciens-mediated transformation. Transgenic tobacco plants carrying the LTB:ST gene are then subjected to GM1-ELISA revealing that the LTB:ST has assembled into pentamers and displays antigenic determinants from both LTB and ST. Protein accumulation of up to 0.05% total soluble protein is detected. Subsequently, mucosal and systemic humoral responses are elicited in mice orally dosed with transgenic tobacco leaves. This has suggested that the plant-derived LTB:ST is immunogenic via the oral route. These findings are critical for the development of a plant-based vaccine capable of eliciting broader protection against ETEC and targeting both LTB and ST. Features of this platform in comparison to transplastomic approaches are discussed. 相似文献
8.
Matsumoto Y Suzuki S Nozoye T Yamakawa T Takashima Y Arakawa T Tsuji N Takaiwa F Hayashi Y 《Transgenic research》2009,18(2):185-192
Cereal crops such as maize and rice are considered attractive for vaccine production and oral delivery. Here, we evaluated
the rice Oryza sativa for production of As16—an antigen protective against the roundworm Ascaris suum. The antigen was produced as a chimeric protein fused with cholera toxin B subunit (CTB), and its expression level in the
endosperm reached 50 μg/g seed. Feeding the transgenic (Tg) rice seeds to mice elicited an As16-specific serum antibody response
when administered in combination with cholera toxin (CT) as the mucosal adjuvant. Although omitting the adjuvant from the
vaccine formulation resulted in failure to develop the specific immune response, subcutaneous booster immunization with bacterially
expressed As16 induced the antibody response, indicating priming capability of the Tg rice. Tg rice/CT-fed mice orally administered
A. suum eggs had a lower lung worm burden than control mice. This suggests that the rice-delivered antigen functions as a prophylactic
edible vaccine for controlling parasitic infection in animals. 相似文献
9.
J.C. Carrero A. Contreras-Rojas B. Sánchez-Hernández P. Petrosyan R.J. Bobes L. Ortiz-Ortiz J.P. Laclette 《Experimental parasitology》2010,126(3):359-365
Entamoeba histolytica antigens recognized by salivary IgA from infected patients include the 29 kDa antigen (Eh29), an alkyl hydroperoxide reductase. Here, we investigate the potential of recombinant Eh29 and an Eh29-cholera toxin subunit B (CTxB) fusion protein to confer protection against intestinal amoebiasis after oral immunization. The purified Eh29-CTxB fusion retained the critical ability to bind ganglioside GM1, as determined by ELISA. Oral immunization of C3H/HeJ mice with Eh29 administered in combination with a subclinical dose of whole cholera toxin, but not as an Eh29-CTxB fusion, induced elevated levels of intestinal IgA and serum IgG anti-Eh29 antibodies that inhibited trophozoites adherence to MDCK cell monolayers. The 80% of immunized mice seen to develop IgA and IgG immune responses showed no evidence of infection in tissue sections harvested following intracecal challenge with virulent E. histolytica trophozoites. These results suggest that Eh29 is capable of inducing protective anti-amoebic immune responses in mice following oral immunization and could be used in the development of oral vaccines against amoebiasis. 相似文献
10.
Guo L Li X Tang F He Y Xing Y Deng X Xi T 《Applied microbiology and biotechnology》2012,93(5):1937-1945
Epitope vaccine based on urease of Helicobacter pylori is a promising option for prophylactic and therapeutic vaccination against H. pylori infection. In this study, we constructed an epitope vaccine with mucosal adjuvant cholera toxin B subunit (CTB) and an epitope
(UreA183-203) of H. pylori urease A subunit named CTB-UA. The CTB-UA fusion protein was expressed in Escherichia coli, and the purified protein was used for intraperitoneal immunization experiments in BALB/c mice. The experimental results
indicated that anti-CTB-UA antibody could recognize both H. pylori urease A subunit (UreA) and urease B subunit (UreB). Besides, the CTB-UA epitope vaccine had good immunogenicity and immunoreactivity
and could induce specific neutralizing antibodies which showed effectively inhibitory effect on the enzymatic activity of
H. pylori urease. CTB-UA is a promising molecule to be investigated as H. pylori vaccine antigen candidate. 相似文献
11.
Expression and immunogenicity of an <Emphasis Type="Italic">Escherichia coli</Emphasis> K99 fimbriae subunit antigen in soybean 总被引:5,自引:0,他引:5
Enterotoxigenic Escherichia coli (ETEC) cause acute diarrhea in humans and farm animals, and can be fatal if the host is left untreated. As a potential alternative to traditional needle vaccination of cattle, we investigated the feasibility of expressing the major K99 fimbrial subunit, FanC, in soybean (Glycine max) for use as an edible subunit vaccine. As a first step in this developmental process, a synthetic version of fanC was optimized for expression in the cytosol and transferred to soybean via Agrobacterium-mediated transformation. Western analysis of T0 events revealed the presence of a peptide with the expected mobility for FanC in transgenic protein extracts, and immunofluorescense confirmed localization to the cytosol. Two T0 lines, which accumulated FanC to levels near 0.5% of total soluble protein, were chosen for further molecular characterization in the T1 and T2 generations. Mice immunized intraperitoneally with protein extract derived from transgenic leaves expressing synthetic FanC developed significant antibody titers against bacterially derived FanC and produced antigen-specific CD4+ T lymphocytes, demonstrating the ability of transgenic FanC to function as an immunogen. These experiments are the first to demonstrate the expression and immunogenicity of a model subunit antigen in the soybean system, and mark the first steps toward the development of a K99 edible vaccine to protect against ETEC. 相似文献
12.
Expression of Escherichia coli heat-labile enterotoxin B subunit (LTB) in Saccharomyces cerevisiae 总被引:1,自引:0,他引:1
Rezaee MA Rezaee A Moazzeni SM Salmanian AH Yasuda Y Tochikubo K Pirayeh SN Arzanlou M 《Journal of microbiology (Seoul, Korea)》2005,43(4):354-360
Heat-labile enterotoxin B subunit (LTB) of enterotoxigenic Escherichia coli (ETEC) is both a strong mucosal adjuvant and immunogen. It is a subunit vaccine candidate to be used against ETEC-induced diarrhea. It has already been expressed in several bacterial and plant systems. In order to construct yeast expressing vector for the LTB protein, the eltB gene encoding LTB was amplified from a human origin enterotoxigenic E. coli DNA by PCR. The expression plasmid pLTB83 was constructed by inserting the eltB gene into the pYES2 shuttle vector immediately downstream of the GAL1 promoter. The recombinant vector was transformed into S. cerevisiae and was then induced by galactose. The LTB protein was detected in the total soluble protein of the yeast by SDS-PAGE analysis. Quantitative ELISA showed that the maximum amount of LTB protein expressed in the yeast was approximately 1.9% of the total soluble protein. Immunoblotting analysis showed the yeast-derived LTB protein was antigenically indistinguishable from bacterial LTB protein. Since the whole-recombinant yeast has been introduced as a new vaccine formulation the expression of LTB in S. cerevisiae can offer an inexpensive yet effective strategy to protect against ETEC, especially in developing countries where it is needed most. 相似文献
13.
免疫霍乱毒素B亚单位(CTB)或肠毒素大肠杆菌(ETEC)定居因子CS3可使人体对ETEC的侵染有保护作用.为探索研制ETEC双组分亚单位疫苗的可行性,利用大肠杆菌诱导表达系统表达了CTB与CS3的融合蛋白(CTB/CS3).蛋白质印迹结果表明,诱导表达的29 ku蛋白具有CTB和CS3蛋白双重抗原性.经Ni-NTA亲和层析纯化获得重组蛋白CTB/CS3,复性的重组蛋白可以部分形成五聚体并保留了与神经节苷脂GM1的结合能力.动物实验表明,融合蛋白CTB/CS3具有CTB和CS3蛋白的双重免疫原性,同时,CTB的免疫载体作用提高了CS3的免疫强度. 相似文献
14.
15.
Dania O. Govea-Alonso Marlene A. Tello-Olea Josué Beltrán-López Elizabeth Monreal-Escalante Jorge A. Salazar-Gonzalez Bernardo Bañuelos-Hernández Sergio Rosales-Mendoza 《Molecular biotechnology》2017,59(11-12):482-489
Atherosclerosis is a pathology leading to cardiovascular diseases with high epidemiologic impact; thus, new therapies are required to fight this global health issue. Immunotherapy is a feasible approach to treat atherosclerosis and given that genetically engineered plants are attractive hosts for vaccine development; we previously proved that the plant cell is able to synthesize a chimeric protein called CTB:p210:CETPe, which is composed of the cholera toxin B subunit (CTB) as immunogenic carrier and target epitopes from the cholesteryl ester transfer protein (CETP461–476) and apolipoprotein B100 (p210). Since CTB:p210:CETPe was expressed in tobacco at sufficient levels to evoke humoral responses in mice, its expression in carrot was explored in the present study looking to develop a vaccine in a safe host amenable for oral delivery; avoiding the purification requirement. Carrot cell lines expressing CTB:p210:CETPe were developed, showing accumulation levels up to 6.1 µg/g dry weight. An immunoblot analysis revealed that the carrot-made protein is antigenic and an oral mice immunization scheme led to evidence on the immunogenic activity of this protein; revealing its capability of inducing serum IgG responses against p210 and CETP epitopes. This study represents a step forward in the development of an attractive oral low-cost vaccine to treat atherosclerosis. 相似文献
16.
17.
Sharma MK Jani D Thungapathra M Gautam JK Meena LS Singh Y Ghosh A Tyagi AK Sharma AK 《Journal of biotechnology》2008,135(1):22-27
18.
Infections by the intracellular protozoan parasite Toxoplasma gondii are widely prevalent in humans and other animals which can cause severe or lethal toxoplasmosis. So the development of a more effective vaccine is needed urgently. A multiantigenic vaccine against toxoplasmosis was constructed in the present study, which contains two T. gondii antigens, SAG1 and MIC4 on the basis of previous immunological and immunization studies. The eukaryotic plasmid pcDNA3.1-SAG1-MIC4, pcDNA3.1-SAG1, pcDNA3.1-MIC4 were constructed first, which can express surface protein SAG1 and microneme protein MIC4 from different stages of T. gondii life cycle, and the expression ability of these DNA vaccine in HeLa cells were examined by Western blot. The efficacy of these plasmids with or without co-administration of a plasmid encoding cholera toxin A2/B as a genetic adjuvant by mucosal way to protect BALB/c mice against toxoplasmosis was evaluated. We found these vaccines were able to elicit a significant humoral and cellular immune response in vaccinated mice and they can increase survival rate and prolong the life of mice that were infected by T. gondii especially in the pcDNA3.1-SAG1-MIC4 group. Co-delivery of cholera toxin A2/B further enhanced the potency of multiantigenic DNA vaccine by intranasal route. These results encourage further research towards achieving vaccinal protection against the T. gondii in animals and humans. 相似文献
19.
The non-toxic B subunit (CT-B) of cholera toxin from Vibrio cholerae is a strong immunogen and amplifies the immune reaction to conjugated antigens. In this work, a synthetic gene encoding for
CT-B was expressed under control of a γ-zein promoter in maize seeds. Levels of CT-B in maize plants were determined via ganglioside
dependent ELISA. The highest expression level recorded in T1 generation seeds was 0.0014% of total aqueous soluble protein (TASP). Expression level of the same event in the T2 generation was significantly increased to 0.0197% of TASP. Immunogenicity of maize derived CT-B was evaluated in mice with
an oral immunization trial. Anti-CTB IgG and anti-CTB IgA were detected in the sera and fecal samples of the orally immunized
mice, respectively. The mice were protected against holotoxin challenge with CT. An additional group of mice was administrated
with an equal amount (5 μg per dose each) of mixed maize-derived CT-B and LT-B (B subunit of E. coli heat labile toxin). In the sera and fecal samples obtained from this group, the specific antibody levels were enhanced compared
to either the same or a higher amount of CT-B alone. These results suggest that a synergistic action may be achieved using
a CT-B and LT-B mixture that can lead to a more efficacious combined vaccine to target diarrhea induced by both cholera and
enterotoxigenic strains of Escherichia coli. 相似文献