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John Hwa Lee - One of the best experts on this subject based on the ideXlab platform.
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salmonella gallinarum delivering m2ecd40l in protein and dna formats acts as a bivalent vaccine against Fowl Typhoid and h9n2 infection in chickens
Veterinary Research, 2018Co-Authors: Irshad Ahmed Hajam, Je Hyoung Kim, John Hwa LeeAbstract:Fowl Typhoid (FT), a septicemic disease caused by Salmonella Gallinarum (SG), and H9N2 influenza infection are two economically important diseases that affect poultry industry worldwide. Herein, we exploited a live attenuated SG mutant (JOL967) to deliver highly conserved extracellular domains of H9N2 M2 (M2e) to induce protective immunity against both H9N2 infection and FT. To increase the immunogenicity of M2e, we physically linked it with CD40L and cloned the fusion gene into either prokaryotic constitutive expression vector pJHL65 or mammalian expression vector pcDNA3.1+. Then pJHL65-M2eCD40L or pcDNA-M2eCD40L recombinant plasmid was electroporated into JOL967 strain and the resultant clones were designated as JOL2074 and JOL2076, respectively. We demonstrated that the chickens vaccinated once orally with a co-mix of JOL2074 and JOL2076 strains elicited significantly (p < 0.05) higher M2e-specific humoral and cell-mediated immunity compared to JOL2074 alone vaccinated group. However, SG-specific immune responses were comparable in both the vaccination groups. On challenge with the virulent H9N2 virus (105 TCID50) at 28th day post-vaccination, chickens that received a co-mix of JOL2074 plus JOL2076 strains exhibited significantly (p < 0.05) lower lung inflammation and viral load in both lungs and cloacal samples than JOL2074 alone vaccinated group. Against challenge with the lethal wild-type SG, both the vaccination groups exhibited only 12.5% mortality compared to 75% mortality observed in the control group. In conclusion, we show that SG delivering M2eCD40L can act as a bivalent vaccine against FT and H9N2 infection and further studies are warranted to develop this SG-M2eCD40L vaccine as a broadly protective vaccine against avian influenza virus subtypes.
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oral immunization with a novel attenuated salmonella gallinarum encoding infectious bronchitis virus spike protein induces protective immune responses against Fowl Typhoid and infectious bronchitis in chickens
Veterinary Research, 2018Co-Authors: Irshad Ahmed Hajam, Je Hyoung Kim, John Hwa LeeAbstract:Fowl Typhoid (FT), a septicemic disease caused by Salmonella Gallinarum (SG), and infectious bronchitis (IB) are two economically important avian diseases that affect poultry industry worldwide. Herein, we exploited a live attenuated SG mutant, JOL967, to deliver spike (S) protein 1 of IB virus (V) to elicit protective immunity against both FT and IB in chickens. The codon optimized S1 nucleotide sequence was cloned in-frame into a prokaryotic constitutive expression vector, pJHL65. Subsequently, empty pJHL65 or recombinant pJHL65-S1 plasmid was electroporated into JOL967 and the resultant clones were designated as JOL2068 and JOL2077, respectively. Our results demonstrated that the chickens vaccinated once orally with JOL2077 elicited significantly (p < 0.05) higher IBV-specific humoral and cell-mediated immunity compared to JOL2068 and PBS control groups. Consequently, on challenge with the virulent IBV strain at 28th day post-vaccination, JOL2077 vaccinated birds displayed significantly (p < 0.05) lower inflammatory lesions in virus-targeted tissues compared to control groups. Furthermore, 33.3% (2 of 6) of birds vaccinated with JOL2077 vaccine had shown virus recovery from tracheal tissues compared to 100% (6 of 6) recovery obtained in both the control groups. Against wild-type SG lethal challenge, both JOL2077 and JOL2068 vaccinated groups exhibited only 10% mortality compared to 80% mortality observed in PBS control group. In conclusion, we show that JOL2077 can induce efficient IBV- and carrier-specific protective immunity and can act as a bivalent vaccine against FT and IB. Further studies are warranted to investigate the potential of JOL2077 vaccine in broiler and young layer birds.
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incorporation of membrane anchored flagellin into salmonella gallinarum bacterial ghosts induces early immune responses and protection against Fowl Typhoid in young layer chickens
Veterinary Immunology and Immunopathology, 2018Co-Authors: Irshad Ahmed Hajam, Je Hyoung Kim, John Hwa LeeAbstract:Abstract The present study aimed to investigate whether the incorporation of flagellin, a TLR5 agonist, in the bacterial ghosts (BGs) of Salmonella Gallinarum can enhance protective immune responses against Fowl Typhoid, a septicemic disease of poultry, in chickens. BGs are empty cell envelopes derived from Gram-negative bacteria through the bacteriophage phiX174 gene E mediated lysis. In this study, the S. Gallinarum ghosts carrying flagellin were genetically constructed utilizing a lysis plasmid pJHL184-flagellin, designed for the coexpression of the flagellin and the lysis protein E. The adjuvant effect of flagellin was evaluated by immunizing seven day old brown nick layer chicks once orally with either S. Gallinarum-flagellin (SG-fliC) ghosts or S. Gallinarum (SG) ghosts alone. Our results showed that immunization with the SG-fliC ghosts elicited early and higher systemic (IgG) and mucosal (IgA) antibody responses compared to the SG ghosts alone, although not always statistically significant. Flow cytometric analysis of the CD3 + CD4+ and the CD3 + CD8+ T cell populations in peripheral blood mononuclear cells were higher in chickens immunized with the SG-fliC ghosts compared to the chickens vaccinated with the SG ghosts alone. Furthermore, the chickens immunized with SG-fliC ghosts exhibited significantly (p
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Intracellular delivery of HA1 subunit antigen through attenuated Salmonella Gallinarum act as a bivalent vaccine against Fowl Typhoid and low pathogenic H5N3 virus
Veterinary Research, 2017Co-Authors: Nitin Machindra Kamble, Kim Je Hyoung, John Hwa LeeAbstract:AbstractIntroduction of novel inactivated oil-emulsion vaccines against different strains of prevailing and emerging low pathogenic avian influenza (LPAI) viruses is not an economically viable option for poultry. Engineering attenuated Salmonella Gallinarum (S. Gallinarum) vaccine delivering H5 LPAI antigens can be employed as a bivalent vaccine against Fowl Typhoid and LPAI viruses, while still offering economic viability and sero-surveillance capacity. In this study, we developed a JOL1814 bivalent vaccine candidate against LPAI virus infection and Fowl Typhoid by engineering the attenuated S. Gallinarum to deliver the globular head (HA1) domain of hemagglutinin protein from H5 LPAI virus through pMMP65 constitutive expression plasmid. The important feature of the developed JOL1814 was the delivery of the HA1 antigen to cytosol of peritoneal macrophages. Immunization of chickens with JOL1814 produced significant level of humoral, mucosal, cellular and IL-2, IL-4, IL-17 and IFN-γ cytokine immune response against H5 HA1 and S. Gallinarum antigens in the immunized chickens. Post-challenge, only the JOL1814 immunized chicken showed significantly faster clearance of H5N3 virus in oropharyngeal and cloacal swabs, and 90% survival rate against lethal challenge with a wild type S. Gallinarum. Furthermore, the JOL1814 immunized were differentiated from the H5N3 LPAI virus infected chickens by matrix (M2) gene-specific real-time PCR. In conclusion, the data from the present showed that the JOL1814 can be an effective bivalent vaccine candidate against H5N3 LPAI and Fowl Typhoid infection in poultry while still offering sero-surveillance property against H5 avian influenza virus.
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evaluation of salmonella gallinarum ghost formulated with montanide isa 70 vg adjuvant as a vaccine against Fowl Typhoid
Acta Veterinaria Hungarica, 2015Co-Authors: Chetan V Jawale, Sang-youel Park, Nithiphonh Somsanith, John Hwa LeeAbstract:Escherichia coli heat-labile enterotoxin B subunit (LTB) protein is a potent adjuvant. Salmonella Gallinarum ghosts carrying LTB (S. Gallinarum-LTB ghosts) were genetically constructed using a plasmid, pJHL187-LTB, designed for the co-expression of the LTB and E lysis proteins. This study evaluates the immunopotentiating effects of Montanide™ ISA 70 VG on S. Gallinarum-LTB ghost vaccination against Fowl Typhoid. Five-week-old layer chickens were injected intramuscularly with sterile PBS (non-immunised control, Group A), S. Gallinarum-LTB ghost (Group B) or S. Gallinarum-LTB ghost emulsified with Montanide™ ISA 70 VG adjuvant (Group C). Chickens from both Groups B and C showed significant induction of antigen-specific systemic IgG response compared to controls; in addition, Group C showed enhanced induction of systemic IgG response compared to Group B. We observed significant induction of antigen-specific lymphocyte proliferative response and increased mRNA levels of Th1 cytokines (IFN-γ and IL2) in both Groups B and C. Furthermore, in the challenge experiment with a virulent strain of S. Gallinarum, Group C showed higher survival rates compared with other groups. These results indicate that vaccination with the S. Gallinarum-LTB ghost in combination with Montanide™ ISA 70 VG may enhance the protective immunity against Fowl Typhoid.
Atul A Chaudhari - One of the best experts on this subject based on the ideXlab platform.
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generation of a safety enhanced salmonella gallinarum ghost using antibiotic resistance free plasmid and its potential as an effective inactivated vaccine candidate against Fowl Typhoid
Vaccine, 2014Co-Authors: Chetan V Jawale, Atul A ChaudhariAbstract:Abstract A safety enhanced Salmonella Gallinarum (SG) ghost was constructed using an antibiotic resistance gene free plasmid and evaluated its potential as Fowl Typhoid (FT) vaccine candidate. The antibiotic resistance free pYA3342 plasmid possesses aspartate semialdehyde dehydrogenase gene which is complimentary to the deletion of the chromosomal asd gene in the bacterial host. This plasmid was incorporated with a ghost cassette containing the bacteriophage PhiX174 lysis gene E , designated as pJHL101. The plasmid pJHL101 was transformed into a two virulence genes - deleted SG. The SG ghosts with tunnel formation and loss of cytoplasmic contents were observed by scanning electron microscopy and transmission electron microscopy. The cell viability of the culture solution was decreased to 0% at 24 h after the induction of gene E expression by an increase in temperature from 37 °C to 42 °C. The safety and protective efficacy of the SG ghost vaccine was further examined in chickens which were divided into three groups: group A (non-immunized control), group B (orally immunized), and group C (intramuscularly immunized). The birds were immunized at 7 d of age. No clinical symptoms associated with FT such as anorexia, depression and greenish diarrhea were observed in the immunized chickens. Upon challenge with a virulent SG strain at 3 week post-immunization, the chickens immunized with the SG ghost via various routes were efficiently protected, as shown by significantly lower mortality and post-mortem lesions in comparison with control group. In addition, all the immunized chickens showed significantly higher antibody responses accompanied by a potent antigen-specific lymphocyte proliferative response along with significantly increased numbers of CD4 + and CD8 + T lymphocytes. Overall, our results provide a promising approach of generating SG ghosts using the antibiotic resistance free plasmid in order to prepare a non-living bacterial vaccine candidate which could be environmentally safe yet efficient to prevent FT in chickens.
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construction of a salmonella gallinarum ghost as a novel inactivated vaccine candidate and its protective efficacy against Fowl Typhoid in chickens
Veterinary Research, 2012Co-Authors: Atul A Chaudhari, Chetan V JawaleAbstract:In order to develop a novel, safe and immunogenic Fowl Typhoid (FT) vaccine candidate, a Salmonella Gallinarum ghost with controlled expression of the bacteriophage PhiX174 lysis gene E was constructed using pMMP99 plasmid in this study. The formation of the Salmonella Gallinarum ghost with tunnel formation and loss of cytoplasmic contents was observed by scanning electron microscopy and transmission electron microscopy. No viable cells were detectable 24 h after the induction of gene E expression by an increase in temperature from 37 °C to 42 °C. The safety and protective efficacy of the Salmonella Gallinarum ghost vaccine was tested in chickens that were divided into four groups: group A (non-immunized control), group B (orally immunized), group C (subcutaneously immunized) and group D (intramuscularly immunized). The birds were immunized at day 7 of age. None of the immunized animals showed any adverse reactions such as abnormal behavior, mortality, or signs of FT such as anorexia, depression, or diarrhea. These birds were subsequently challenged with a virulent Salmonella Gallinarum strain at 3 weeks post-immunization (wpi). Significant protection against the virulent challenge was observed in all immunized groups based on mortality and post-mortem lesions compared to the non-immunized control group. In addition, immunization with the Salmonella Gallinarum ghosts induced significantly high systemic IgG response in all immunized groups. Among the groups, orally-vaccinated group B showed significantly higher levels of secreted IgA. A potent antigen-specific lymphocyte activation response along with significantly increased percentages of CD4+ and CD8+ T lymphocytes found in all immunized groups clearly indicate the induction of cellular immune responses. Overall, these findings suggest that the newly constructed Salmonella Gallinarum ghost appears to be a safe, highly immunogenic, and efficient non-living bacterial vaccine candidate that protects against FT.
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safety and efficacy of a virulence gene deleted live vaccine candidate for Fowl Typhoid in young chickens
Avian Pathology, 2011Co-Authors: Kiku Matsuda, Atul A Chaudhari, John Hwa LeeAbstract:The safety and efficacy of a live lon-and-cpxR-deleted Salmonella enterica serovar Gallinarum (SG) vaccine candidate (JOL916) was evaluated in young layer chickens. Vaccinated (n=25) and unvaccinated (n=25) groups were organized, respectively, at 1, 2, 3, and 4 weeks of age. One-week-old and 2-week-old chickens were orally inoculated with 2×10(7) colony-forming units of JOL916, and orally challenged with 2 x 10(6) colony-forming units of a wild-type SG strain at the third week post vaccination (w.p.v.). Doses of vaccination and challenge were increased 10-fold for 3-week-old and 4-week-old chickens. SG-antigen-specific peripheral lymphocyte proliferation response and concentrations of plasma IgG and secretary IgA in the intestine were examined at the second w.p.v. Gross lesions of the liver and spleen and recovery of the vaccine strain from the spleen were also examined at the second w.p.v. No evidence of side effects was detected by observation of general condition and body weight gain in all vaccinated groups. No, or very mild, gross lesions in the chickens were observed in the liver and/or spleen after vaccination. Significant cellular immune responses and systemic IgG responses were induced after vaccination in all age groups. Elevation of secretary IgA concentration was significant in the group, vaccinated at the age of 1 week. Depression scores after challenge were significantly lower in the vaccinated groups, as compared with the corresponding control groups. Significant reductions of death rates were observed in all vaccinated groups, as compared with the equivalent unvaccinated groups. Thus, the oral vaccination of young chickens with JOL916 was demonstrated to be safe. Moreover, it offered efficient protection against Fowl Typhoid.
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safety evaluation and immunogenicity of arabinose based conditional lethal salmonella gallinarum mutant unable to survive ex vivo as a vaccine candidate for protection against Fowl Typhoid
Avian Diseases, 2011Co-Authors: Atul A Chaudhari, Kiku MatsudaAbstract:Abstract In seeking to develop a safe Fowl Typhoid (FT) vaccine, a novel candidate lacking cpxR, lon, and asd Salmonella Gallinarum (SG) genes was constructed with the plasmid-containing araC::ParaBAD::asd system. A balanced-lethal host-vector system based on the essential bacterial gene for aspartate β-semialdehyde dehydrogenase (asd) was used to construct the SG mutant strain. A plasmid (p15A ori) with an araC::ParaBAD::asd cassette was introduced into an auxotrophic mutant to prevent ex vivo survival. The safety, immunity, and protective properties of the SG mutant were evaluated. Inoculation of the mutant at 106 colony-forming units (CFU) did not result in recovery in feces and internal organs, whereas inoculation at 108 and 1010 CFU resulted in moderate bacterial recovery from feces and organs. Birds immunized with the mutant were challenged with a virulent SG strain at day 14 postimmunization; significantly reduced mortality and induced plasma immunoglobulin (Ig)G and mucosal IgA responses were note...
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evaluation of safety and protection efficacy on cpxr and lon deleted mutant of salmonella gallinarum as a live vaccine candidate for Fowl Typhoid
Vaccine, 2011Co-Authors: Kiku Matsuda, Atul A ChaudhariAbstract:We evaluated a recently developed live Fowl Typhoid (FT) vaccine candidate, JOL916, the cpxR/lon mutant of Salmonella Gallinarum (SG), for safety and protection efficacy in 5-week-old layer chickens. Intramuscular vaccination with JOL916 revealed no or very few lesions in livers and spleens of the animals until the fourth week post-vaccination (wpv). This candidate clearly induced cellular immune responses in 5 of 5 chickens on the first and second wpv based on the peripheral lymphocyte proliferation assay. Systemic IgG responses were observed in 5 of 5 chickens from the first wpv and dramatic elevations were observed on the second and third wpv. Vaccination of chickens offered efficient protection against challenge by a wild-type SG; only slight anorexia and depression were temporarily observed after challenge in the vaccinated group while 100% mortality was observed in the positive control group. Body weight increases per day were slightly reduced between the 3rd and 6th day post challenge (dpc) compared to the negative control group; it was recovered from the 6th dpc. Collectively, these results demonstrate the safety and protective efficacy of JOL916 as a live vaccine for systemic FT.
Hyukjoon Kwon - One of the best experts on this subject based on the ideXlab platform.
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optimized detoxification of a live attenuated vaccine strain sg9r to improve vaccine strategy against Fowl Typhoid
Vaccine, 2021Co-Authors: Namhyung Kim, Sunmin Ahn, Kangseuk Choi, Hyukjoon KwonAbstract:The live attenuated vaccine strain, SG9R, has been used against Fowl Typhoid worldwide, but it can revert to the pathogenic smooth strain owing to single nucleotide changes such as nonsense mutations in the rfaJ gene. As SG9R possesses an intact Salmonella plasmid with virulence genes, it exhibits dormant pathogenicity and can cause Fowl Typhoid in young chicks and stressed or immunocompromised brown egg-laying hens. To tackle these issues, we knocked out the rfaJ gene of SG9R (named Safe-9R) to eliminate the reversion risk and generated detoxified strains of Safe-9R by knocking out lpxL, lpxM, pagP, and phoP/phoQ genes to attenuate the virulence. Among the knockout strains, live ΔlpxL- (Dtx-9RL) and ΔlpxM-9R (Dtx-9RM) strains induced remarkably less expression of inflammatory cytokines in chicken macrophage cells, and oil emulsion (OE) Dtx-9RL did not cause body weight loss in chicks. Live Dtx-9RM exhibited efficacy against field strain challenge in one week without any bacterial re-isolation, while the un-detoxified strains showed the development of severe liver lesions and re-isolation of challenged strains. Thus, SG9R was optimally detoxified by knockout of lpxL and lpxM, and Dtx-9RL and Dtx-9RM might be applicable as OE and live vaccines, respectively, to prevent Fowl Typhoid irrespective of the age of chickens.
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establishment of a live vaccine strain against Fowl Typhoid and paraTyphoid
Korean Journal of Veterinary Research, 2015Co-Authors: Sunhee Cho, Sunjoong Kim, Youngjin Ahn, Taeeun Kim, Won Huh, Youngsik Moon, Byunghyung Lee, Jaehong Kim, Hyukjoon KwonAbstract:To develop a live vaccine strain against Fowl Typhoid and paraTyphoid caused by Salmonella serovar Gallinarum biovar Gallinarum (Salmonella Gallinarum) and Salmonella serovar Enteritidis (Salmonella Enteritidis), respectively, several nalidixic acid resistant mutants were selected from lipopolysaccharide (LPS) rough strains of Salmonella Gallinarum that escaped from fatal infection of a LPS-binding lytic bacteriophage. A non-virulent and immunogenic vaccine strain of Salmonella Gallinarum, SR2-N6, was established through in vivo pathogenicity and protection efficacy tests. SR2-N6 was highly protective against Salmonella Gallinarum and Salmonella Enteritidis and safer than Salmonella Gallinarum vaccine strain SG 9R in the condition of protein-energy malnutrition. Thus, SR2- N6 may be a safe and efficacious vaccine strain to prevent both Fowl Typhoid and paraTyphoid.
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pathogenicity of sg 9r a rough vaccine strain against Fowl Typhoid
Vaccine, 2011Co-Authors: Hyukjoon Kwon, Sunhee ChoAbstract:Abstract SG 9R, a rough vaccine strain of Salmonella gallinarum , has been used for the prevention of Fowl Typhoid and paraTyphoid in the world despite the presence of residual virulence. SG 9R-like rough strains have been recently isolated from Fowl Typhoid cases; however, molecular markers to differentiate SG 9R from field strains are not well-characterized and the molecular mechanisms of SG 9R residual virulence are unclear. Therefore, we analyzed LPS biosynthesis ( rfa gene cluster) and virulence genes ( spv , SPI-2) of both SG 9R and S. gallinarum rough field strains. SG 9R carried a unique nonsense mutation in rfaJ (TCA to TAA) and a shared rfaZ mutation (G-deletion) by rough and smooth S. gallinarum strains. SG 9R also carried intact SPI-2 and spvC , B , A , and R (except deleted spvD ). SG 9R-like rough strains ( n = 10) carried identical mutations in virulence-related genes to SG 9R. SG 9R and SG 9R-like rough strains did not demonstrate significant mortality or liver lesions under normal conditions. However, Fowl Typhoid was successfully reproduced in the present study by SG 9R inoculation to 1-day-old male brown layer chicks per os following starvation. Therefore, the LPS defect may be one of the major mechanisms of SG 9R attenuation, and the possession of intact SPI-2, spvC , B , A , and R virulence genes may be associated with residual SG 9R virulence.
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characterization of a t7 like lytic bacteriophage φsg jl2 of salmonella enterica serovar gallinarum biovar gallinarum
Applied and Environmental Microbiology, 2008Co-Authors: Hyukjoon Kwon, Jihye Jeong, Se Chang Park, Yong Ho ParkAbstract:T7-like viruses have short noncontractile tails and are members of the family Podoviridae. To date, eight strains have been assigned as Enterobacteria phage T7 and three strains (T3, T7, and φYeO3-12) have been characterized genomically (http://www.ncbi.nlm.nih.gov/ICTVdb/Ictv/index.htm) (19, 50, 51). Genetic recombination between T7-like viruses infecting different bacterial genera or different species has been demonstrated, and T3 may have evolved from an ancient phage generated by recombination between yersiniophages φA1122 and φYeO3-12 (20, 51). Horizontal genetic transfer results in genomic mosaicism of phages, which hinders their hierarchical classification (22, 37). However, common genetic components and layouts observed among T7-like viruses may support the idea that they crossed a “Darwinian threshold” and have been undergoing vertical evolution (26, 79). Therefore, they may be useful in understanding genetic variations of closely related T7-like phages during host adaptation. However, current genomic data are insufficient to permit such detailed analysis. Additional genome sequences of closely related T7-like viruses are required to gain insight into their vertical evolution. Fowl Typhoid is an acute septicemic disease occurring in adult chickens. The disease is characterized by anemia, leukocytosis, and hemorrhage and is an economically disastrous disease in the poultry industry (53). The causative agent, Salmonella enterica serovar Gallinarum biovar Gallinarum, is classified in serogroup D and is both nonmotile and host adapted (3, 53). Differentiation of serovar Gallinarum biovar Gallinarum from frequent avian serogroup D Salmonella strains, such as S. enterica serovar Gallinarum biovar Pullorum and S. enterica serovar Enteritidis, has been partially successful (33, 34), and differentiation of field strains of serovar Gallinarum biovar Gallinarum from the rough vaccine strain SG-9R has become important because of nationwide vaccination in some countries. The appearance of multidrug-resistant serovar Gallinarum biovar Gallinarum strains in the field has prompted increasing concerns about phage therapy, similar to other bacterial diseases (5, 30, 35, 64, 68), but candidate phages that are lytic to broad ranges of serovar Gallinarum biovar Gallinarum strains have never been reported. Fowl Typhoid has been reported to spread via the fecal-oral route, but recently, Fowl Typhoid was reproduced by intratracheal challenge with serovar Gallinarum biovar Gallinarum (4). In this study, we report the basic biological properties and complete genomic sequence of a new Salmonella T7-like virus, φSG-JL2. It is lytic to serovar Gallinarum biovar Gallinarum and has a double-stranded DNA of 38,815 bp with 55 putative genes. Comparative genomic analyses have demonstrated the close relationships of φSG-JL2 with φYeO3-12 from Yersinia enterocolitica O3 and with T3 from Escherichia coli and have provided molecular clues to understand host adaptations of related phages. The obligate specificity and broad lytic activity of φSG-JL2 may be useful for differentiation of serovar Gallinarum biovar Gallinarum from S. enterica serovar Enteritidis and serovar Gallinarum biovar Pullorum, and the prophylactic efficacy of φSG-JL2 against Fowl Typhoid was tested with a respiratory model of Fowl Typhoid.
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characterization of a t7 like lytic bacteriophage φsg jl2 of salmonella enterica serovar gallinarum biovar gallinarum
Applied and Environmental Microbiology, 2008Co-Authors: Hyukjoon Kwon, Yong Ho Park, Jihye Jeong, Se Chang Park, Han Sang Yoo, Sunhee Cho, Taeeun Kim, Jaehong Kim, Yongjin Won, Sunjoong KimAbstract:PhiSG-JL2 is a newly discovered lytic bacteriophage infecting Salmonella enterica serovar Gallinarum biovar Gallinarum but is nonlytic to a rough vaccine strain of serovar Gallinarum biovar Gallinarum (SG-9R), S. enterica serovar Enteritidis, S. enterica serovar Typhimurium, and S. enterica serovar Gallinarum biovar Pullorum. The phiSG-JL2 genome is 38,815 bp in length (GC content, 50.9%; 230-bp-long direct terminal repeats), and 55 putative genes may be transcribed from the same strand. Functions were assigned to 30 genes based on high amino acid similarity to known proteins. Most of the expected proteins except tail fiber (31.9%) and the overall organization of the genomes were similar to those of yersiniophage phiYeO3-12. phiSG-JL2 could be classified as a new T7-like virus and represents the first serovar Gallinarum biovar Gallinarum phage genome to be sequenced. On the basis of intraspecific ratios of nonsynonymous to synonymous nucleotide changes (Pi[a]/Pi[s]), gene 2 encoding the host RNA polymerase inhibitor displayed Darwinian positive selection. Pretreatment of chickens with phiSG-JL2 before intratracheal challenge with wild-type serovar Gallinarum biovar Gallinarum protected most birds from Fowl Typhoid. Therefore, phiSG-JL2 may be useful for the differentiation of serovar Gallinarum biovar Gallinarum from other Salmonella serotypes, prophylactic application in Fowl Typhoid control, and understanding of the vertical evolution of T7-like viruses.
Paul Barrow - One of the best experts on this subject based on the ideXlab platform.
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evaluation of pathogenicity of salmonella gallinarum strains harbouring deletions in genes whose orthologues are conserved pseudogenes in s pullorum
PLOS ONE, 2018Co-Authors: Diego Felipe Alves Batista, Adriana Maria De Almeida, Paul Barrow, Oliveiro Caetano Freitas Neto, Grazieli Maboni, Tatiane Furtado De Carvalho, Thaynara Parente De Carvalho, Angelo BerchieriAbstract:: The diseases caused by Salmonella Gallinarum and S. Pullorum in chickens known as Fowl Typhoid and pullorum disease, respectively, pose a great threat to the poultry industry mainly in developing countries, since they have already been controlled in the developed ones. These bacteria are very similar at the genomic level but develop distinct host-pathogen relationships with chickens. Therefore, a deep understanding of the molecular mechanisms whereby S. Gallinarum and S. Pullorum interact with the host could lead to the development of new approaches to control and, perhaps, eradicate both diseases from the chicken flocks worldwide. Based on our previous study, it was hypothesised that metabolism-related pseudogenes, fixed in S. Pullorum genomes, could play a role in the distinct host-pathogen interaction with susceptible chickens. To test this idea, three genes (idnT, idnO and ccmH) of S. Gallinarum str. 287/91, which are pseudogenes on the S. Pullorum chromosomes, were inactivated by mutations. These genetically engineered strains grew well on the solid media without any colony morphology difference. In addition, similar growth curves were obtained by cultivation in M9 minimal medium containing D-gluconate as the sole carbon source. Infection of chickens with idnTO mutants led to increased numbers of bacteria in the livers and spleens at 5 days post-infection, but with slightly decreased heterophil infiltration in the spleens when compared to the wild-type strain. On the other hand, no significant phenotypic change was caused by mutation to ccmH genes. Apart from the above-mentioned alterations, all S. Gallinarum strains provoked similar infections, since mortality, clinical signs, macroscopic alterations and immune response were similar to the infected chickens. Therefore, according to the model applied to this study, mutation to the idnTO and ccmH genes showed minor impact on the Fowl Typhoid pathogenesis and so they may be relics from the ancestor genome. Our data hints at a more complex mechanism driving the distinct host-pathogen interaction of S. Gallinarum/Pullorum with chickens than differential inactivation of a few genes.
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inactivation of phopq genes attenuates salmonella gallinarum biovar gallinarum to susceptible chickens
Brazilian Journal of Microbiology, 2018Co-Authors: Lucas Bocchini Rodrigues Alves, Diego Felipe Alves Batista, Fernanda O Barbosa, Marcela Da Silva Rubio, Andrei Itajahy Secundo De Souza, Adriana Maria De Almeida, Paul Barrow, Oliveiro Caetano Freitas Neto, Angelo BerchieriAbstract:Abstract Salmonella Gallinarum is a host-restrict pathogen that causes Fowl Typhoid, a severe systemic disease that is one of the major concerns to the poultry industry worldwide. When infecting the bird, SG makes use of evasion mechanisms to survive and to replicate within macrophages. In this context, phoPQ genes encode a two-component regulatory system (PhoPQ) that regulates virulence genes responsible for adaptation of Salmonella spp. to antimicrobial factors such as low pH, antimicrobial peptides and deprivation of bivalent cations. The role of the mentioned genes to SG remains to be investigated. In the present study a phoPQ-depleted SG strain (SG ΔphoPQ) was constructed and its virulence assessed in twenty-day-old laying hens susceptible to Fowl Typhoid. SG ΔphoPQ did cause neither clinical signs nor mortality in birds orally challenged, being non-pathogenic. Furthermore, this strain was not recovered from livers or spleens. On the other hand, chickens challenged subcutaneously with the mutant strain had discreet to moderate pathological changes and also low bacterial counts in liver and spleen tissues. These findings show that SG ΔphoPQ is attenuated to susceptible chickens and suggest that these genes are important during chicken infection by SG.
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identification and characterization of regions of difference between the salmonella gallinarum biovar gallinarum and the salmonella gallinarum biovar pullorum genomes
Infection Genetics and Evolution, 2015Co-Authors: Diego Felipe Alves Batista, Marcos T Oliveira, A.m. De Almeida, Paul Barrow, Oliveiro Caetano Freitas Neto, Antônio Sérgio Ferraudo, Angelo BerchieriAbstract:Salmonella Gallinarum is the causative agent of Fowl Typhoid, a severe septicaemic disease that affects birds of all ages, whereas S. Pullorum causes pullorum disease, a systemic disorder affecting primarily young birds. A proportion of birds with pullorum disease become carriers and are thereby able to transmit S. Pullorum vertically. Although these two pathogens cause distinct diseases, they are otherwise phenotypically and genetically similar. Therefore, the small variations that lead to the differences in virulence must have a genetic basis which currently is unknown. In the present study, we compared the genome sequences of S. Gallinarum (strains: SG287/91 and SG9) and S. Pullorum (strains: SP_CDC, SP_RKS, SP_FCAV, SP_S06) and identified 223 regions of difference (RODs), characterized by indels which were detected by using the software Artemis Comparison Tool. Some of the RODs led to pseudogenes frequently formed by frameshifts and premature stop codons in genes primarily involved in virulence and metabolism. We further verified the presence of some conserved RODs by PCR in 26 isolates of S. Gallinarum and 17 of S. Pullorum in order to extrapolate data analyses from genome comparison to field strains. The variations observed in virulence-related genes of S. Gallinarum and S. Pullorum appear not to be sufficient to explain the differences between the distinct biology of infection of Fowl Typhoid and pullorum disease. Thus, we suggest that the identified pseudogenes affecting metabolism might play a greater role during infection than previously thought.
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oral infection with the salmonella enterica serovar gallinarum 9r attenuated live vaccine as a model to characterise immunity to Fowl Typhoid in the chicken
BMC Veterinary Research, 2005Co-Authors: Paul Wigley, Scott D Hulme, Claire Powers, Richard K Beal, Adrian Smith, Paul BarrowAbstract:Background Salmonella enterica serovar Gallinarum (S. Gallinarum) is the causative agent of Fowl Typhoid, a severe systemic disease of chickens that results in high mortality amongst infected flocks. Due to its virulence, the immune response to S. Gallinarum is poorly characterised. In this study we have utilised infection by the live attenuated S. Gallinarum 9R vaccine strain in inbred chickens to characterise humoral, cellular and cytokine responses to systemic salmonellosis.
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oral infection with the salmonella enterica serovar gallinarum 9r attenuated live vaccine as a model to characterise immunity to Fowl Typhoid in the chicken
BMC Veterinary Research, 2005Co-Authors: Paul Wigley, Scott D Hulme, Claire Powers, Richard K Beal, Adrian Smith, Paul BarrowAbstract:Background: Salmonella enterica serovar Gallinarum (S. Gallinarum) is the causative agent of Fowl Typhoid, a severe systemic disease of chickens that results in high mortality amongst infected flocks. Due to its virulence, the immune response to S. Gallinarum is poorly characterised. In this study we have utilised infection by the live attenuated S. Gallinarum 9R vaccine strain in inbred chickens to characterise humoral, cellular and cytokine responses to systemic salmonellosis. Results: Infection with 9R results in a mild systemic infection. Bacterial clearance at three weeks post infection coincides with increases in circulating anti-Salmonella antibodies, increased T cell proliferation to Salmonella challenge and increased expression of interferon gamma. These responses peak at four weeks post infection, then decline. Only modest increases of expression of the pro-inflammatory cytokine interleukin-1β were detected early in the infection. Conclusion: Infection of chickens with the 9R vaccine strain induces a mild form of systemic salmonellosis. This induces both cellular and humoral immune responses, which peak soon after bacterial clearance. Unlike enteric-associated Salmonella infections the immune response is not prolonged, reflecting the absence of persistence of Salmonella in the gastrointestinal tract. The findings here indicate that the use of the S. Gallinarum 9R vaccine strain is an effective model to study immunity to systemic salmonellosis in the chicken and may be employed in further studies to determine which components of the immune response are needed for protection.
Chetan V Jawale - One of the best experts on this subject based on the ideXlab platform.
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evaluation of salmonella gallinarum ghost formulated with montanide isa 70 vg adjuvant as a vaccine against Fowl Typhoid
Acta Veterinaria Hungarica, 2015Co-Authors: Chetan V Jawale, Sang-youel Park, Nithiphonh Somsanith, John Hwa LeeAbstract:Escherichia coli heat-labile enterotoxin B subunit (LTB) protein is a potent adjuvant. Salmonella Gallinarum ghosts carrying LTB (S. Gallinarum-LTB ghosts) were genetically constructed using a plasmid, pJHL187-LTB, designed for the co-expression of the LTB and E lysis proteins. This study evaluates the immunopotentiating effects of Montanide™ ISA 70 VG on S. Gallinarum-LTB ghost vaccination against Fowl Typhoid. Five-week-old layer chickens were injected intramuscularly with sterile PBS (non-immunised control, Group A), S. Gallinarum-LTB ghost (Group B) or S. Gallinarum-LTB ghost emulsified with Montanide™ ISA 70 VG adjuvant (Group C). Chickens from both Groups B and C showed significant induction of antigen-specific systemic IgG response compared to controls; in addition, Group C showed enhanced induction of systemic IgG response compared to Group B. We observed significant induction of antigen-specific lymphocyte proliferative response and increased mRNA levels of Th1 cytokines (IFN-γ and IL2) in both Groups B and C. Furthermore, in the challenge experiment with a virulent strain of S. Gallinarum, Group C showed higher survival rates compared with other groups. These results indicate that vaccination with the S. Gallinarum-LTB ghost in combination with Montanide™ ISA 70 VG may enhance the protective immunity against Fowl Typhoid.
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generation of a safety enhanced salmonella gallinarum ghost using antibiotic resistance free plasmid and its potential as an effective inactivated vaccine candidate against Fowl Typhoid
Vaccine, 2014Co-Authors: Chetan V Jawale, Atul A ChaudhariAbstract:Abstract A safety enhanced Salmonella Gallinarum (SG) ghost was constructed using an antibiotic resistance gene free plasmid and evaluated its potential as Fowl Typhoid (FT) vaccine candidate. The antibiotic resistance free pYA3342 plasmid possesses aspartate semialdehyde dehydrogenase gene which is complimentary to the deletion of the chromosomal asd gene in the bacterial host. This plasmid was incorporated with a ghost cassette containing the bacteriophage PhiX174 lysis gene E , designated as pJHL101. The plasmid pJHL101 was transformed into a two virulence genes - deleted SG. The SG ghosts with tunnel formation and loss of cytoplasmic contents were observed by scanning electron microscopy and transmission electron microscopy. The cell viability of the culture solution was decreased to 0% at 24 h after the induction of gene E expression by an increase in temperature from 37 °C to 42 °C. The safety and protective efficacy of the SG ghost vaccine was further examined in chickens which were divided into three groups: group A (non-immunized control), group B (orally immunized), and group C (intramuscularly immunized). The birds were immunized at 7 d of age. No clinical symptoms associated with FT such as anorexia, depression and greenish diarrhea were observed in the immunized chickens. Upon challenge with a virulent SG strain at 3 week post-immunization, the chickens immunized with the SG ghost via various routes were efficiently protected, as shown by significantly lower mortality and post-mortem lesions in comparison with control group. In addition, all the immunized chickens showed significantly higher antibody responses accompanied by a potent antigen-specific lymphocyte proliferative response along with significantly increased numbers of CD4 + and CD8 + T lymphocytes. Overall, our results provide a promising approach of generating SG ghosts using the antibiotic resistance free plasmid in order to prepare a non-living bacterial vaccine candidate which could be environmentally safe yet efficient to prevent FT in chickens.
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attenuated salmonella gallinarum secreting an escherichia coli heat labile enterotoxin b subunit protein as an adjuvant for oral vaccination against Fowl Typhoid
Veterinary Immunology and Immunopathology, 2012Co-Authors: Byung Woo Jeon, Chetan V JawaleAbstract:Abstract In our previous study, we constructed a vaccine candidate (JOL916) for Fowl Typhoid (FT). A live adjuvant Salmonella Gallinarum (SG) strain was generated in the present study to facilitate efficacious oral vaccination with this vaccine. The Escherichia coli eltB gene secreting heat-labile enterotoxin B subunit (LTB) was cloned into an Asd+ plasmid pJHL65. This was transformed into a Δlon ΔcpxR Δasd SG strain and the resulting strain was designated JOL1229. Secretion of LTB from JOL1229 was confirmed with an immunoblot assay. To determine the optimal dose of the strain, 50 six-week-old female chickens were divided into five groups (Groups A–E, n = 10 per group) and orally inoculated with various doses of JOL1229 and JOL916. In Group B (consisting of four parts JOL916 and one part JOL1229), significant cell-mediated immune responses, plasma IgG levels and intestinal secretary IgA levels were induced after inoculation with both strains. On challenge with the wild-type strain, significant reductions in mortality were observed in the group. In addition, after inoculation the LTB strain was not recovered in feces samples, and resulted in no, or very mild, gross lesions in the liver and spleen. Both CD4+ and CD8+ T-cells were significantly increased in peripheral blood samples from the chickens immunized with the LTB strain. Expression of the interleukin-6 (IL-6) gene in splenocytes was induced in the chickens immunized with the LTB strain. These results suggest that oral immunization with the LTB-adjuvant strain, in particular with the four parts JOL916 and one part JOL1229 mixture, increased the immune response and provided efficient protection against FT in chickens.
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construction of a salmonella gallinarum ghost as a novel inactivated vaccine candidate and its protective efficacy against Fowl Typhoid in chickens
Veterinary Research, 2012Co-Authors: Atul A Chaudhari, Chetan V JawaleAbstract:In order to develop a novel, safe and immunogenic Fowl Typhoid (FT) vaccine candidate, a Salmonella Gallinarum ghost with controlled expression of the bacteriophage PhiX174 lysis gene E was constructed using pMMP99 plasmid in this study. The formation of the Salmonella Gallinarum ghost with tunnel formation and loss of cytoplasmic contents was observed by scanning electron microscopy and transmission electron microscopy. No viable cells were detectable 24 h after the induction of gene E expression by an increase in temperature from 37 °C to 42 °C. The safety and protective efficacy of the Salmonella Gallinarum ghost vaccine was tested in chickens that were divided into four groups: group A (non-immunized control), group B (orally immunized), group C (subcutaneously immunized) and group D (intramuscularly immunized). The birds were immunized at day 7 of age. None of the immunized animals showed any adverse reactions such as abnormal behavior, mortality, or signs of FT such as anorexia, depression, or diarrhea. These birds were subsequently challenged with a virulent Salmonella Gallinarum strain at 3 weeks post-immunization (wpi). Significant protection against the virulent challenge was observed in all immunized groups based on mortality and post-mortem lesions compared to the non-immunized control group. In addition, immunization with the Salmonella Gallinarum ghosts induced significantly high systemic IgG response in all immunized groups. Among the groups, orally-vaccinated group B showed significantly higher levels of secreted IgA. A potent antigen-specific lymphocyte activation response along with significantly increased percentages of CD4+ and CD8+ T lymphocytes found in all immunized groups clearly indicate the induction of cellular immune responses. Overall, these findings suggest that the newly constructed Salmonella Gallinarum ghost appears to be a safe, highly immunogenic, and efficient non-living bacterial vaccine candidate that protects against FT.
