Avian pathogenic Escherichia coli (APEC) is a causative agent for a number of extra intestinal diseases and account for significant losses to the poultry industry. Since protective immunity against APEC is largely directed to virulence antigens, we have individually expressed four different viulence antigens, papA, papG, IutA, and CS31A, using an attenuated Salmonella Typhimurium and a plasmid pBB244. Following oral immunization of mice with combination of two or four of these strains, serum IgG and mucosal IgA responses were elicited against each antigen represented in the mixture. The antigen-specific mucosal IgA responses were significantly higher in the group of mice immunized with the heat-labile Escherichia coli enterotoxin B subunit (LTB) strain than those in the group of mice immunized without the LTB strain. While, there was no significant difference between these two groups in antigen-specific serum IgG responses. The results showed that LTB could act as mucosal immune adjuvant. To assess the nature of immunity, the distribution of antigen-specific IgG isotypes was analyzed. All groups promoted Th1-type immunity as determined by the IgG2a/IgG1 ratio. Thus, our findings provided evidence that immunization with a combination of several vaccine strains is one of the strategies of developing effective vaccines against APEC.
Animals ; Enterotoxins ; Escherichia coli ; Immunity, Mucosal* ; Immunization ; Immunoglobulin A ; Immunoglobulin G ; Intestinal Diseases ; Mice* ; Plasmids ; Poultry ; Salmonella typhimurium ; Salmonella Vaccines* ; Salmonella* ; Vaccines ; Virulence

Diarrhea is one of the most common causes of morbidity and mortality in children worldwide. Rotavirus is the most common cause of infectious diarrhea both in developed and developing countries. However, bacterial causes such as Salmonella typhi and Vibrio cholerae still play an important role in developing countries. Newly developed vaccines for rotavirus, S. typhi, and V. choleae are highly immunogenic and safe in children.
Child ; Developing Countries ; Diarrhea ; Humans ; Rotavirus ; Salmonella typhi ; Typhoid-Paratyphoid Vaccines ; Vaccines ; Vibrio cholerae

BACKGROUND: This study is aimed at evaluating immunogenicity by measuring immunoglobulin A (IgA) seroconversion rate through common mucosal immune system and adverse reactions after vaccination of oral live attenuated Salmonella typhi (S. typhi) Ty21a vaccine in Korean population. METHODS: A commercially available oral live attenuated vaccine of S. typhi strain Ty21a (Zerotyph(r) capsule, Boryung Biopharma Co., Seoul, Korea) was given to volunteers; children above 6 years, adolescents, and adults who have never infected with S. typhi nor received S. typhi vaccination. The vaccines were given in three doses, with two day interval between the doses. Seroconversion was determined by ELISPOT (enzyme-linked immunospot) assay. Adverse reactions after vaccination were evaluated in 12 institutions by direct interviewing with vaccinees. RESULTS: A total of 93 volunteers for evaluation of seroconversion were enrolled. Seroconversion rate in the the below 16 year-old group was 73.8% (31/42) and that of over 16 year-old group was 86.3% (44/51), which was not statistically different. Adverse reaction were found in 8.6% (40/465). Gastrointestinal symptoms were most common (6.5%, 30/465). Adverse reactions were found in 5.2% (24/465) after 1st administration, 4.5% (21/462) after 2nd, and 2.6% (12/461) after 3rd. Frequency of adverse reactions was significantly higher after 1st administration (P<0.05). CONCLUSION: Oral live attenuated S. typhi vaccine, Zerotyph(r) capsule, had good immnuogenicity and safety through intestinal immune system.
Adolescent ; Adult ; Child ; Enzyme-Linked Immunospot Assay ; Humans ; Immune System ; Immunoglobulin A ; Salmonella typhi* ; Salmonella* ; Seoul ; Vaccination ; Vaccines ; Volunteers

BACKGROUND: This study is aimed at evaluating immunogenicity by measuring immunoglobulin A (IgA) seroconversion rate through common mucosal immune system and adverse reactions after vaccination of oral live attenuated Salmonella typhi (S. typhi) Ty21a vaccine in Korean population. METHODS: A commercially available oral live attenuated vaccine of S. typhi strain Ty21a (Zerotyph(r) capsule, Boryung Biopharma Co., Seoul, Korea) was given to volunteers; children above 6 years, adolescents, and adults who have never infected with S. typhi nor received S. typhi vaccination. The vaccines were given in three doses, with two day interval between the doses. Seroconversion was determined by ELISPOT (enzyme-linked immunospot) assay. Adverse reactions after vaccination were evaluated in 12 institutions by direct interviewing with vaccinees. RESULTS: A total of 93 volunteers for evaluation of seroconversion were enrolled. Seroconversion rate in the the below 16 year-old group was 73.8% (31/42) and that of over 16 year-old group was 86.3% (44/51), which was not statistically different. Adverse reaction were found in 8.6% (40/465). Gastrointestinal symptoms were most common (6.5%, 30/465). Adverse reactions were found in 5.2% (24/465) after 1st administration, 4.5% (21/462) after 2nd, and 2.6% (12/461) after 3rd. Frequency of adverse reactions was significantly higher after 1st administration (P<0.05). CONCLUSION: Oral live attenuated S. typhi vaccine, Zerotyph(r) capsule, had good immnuogenicity and safety through intestinal immune system.
Adolescent ; Adult ; Child ; Enzyme-Linked Immunospot Assay ; Humans ; Immune System ; Immunoglobulin A ; Salmonella typhi* ; Salmonella* ; Seoul ; Vaccination ; Vaccines ; Volunteers

OBJECTIVETo develop an anti-caries DNA vaccine-loaded Salmonella typhimurium (St) ghost and enhance the efficacy of immune responses induced by anti-caries DNA vaccine via mucosal route.

METHODSBoth pREP4 and PhiX gene E expression plasmids were transformed into StJ357 and then induced with isopropyl β-D-1-thiogalactopyranoside (IPTG). The bacterial ghosts (BG) were collected after wash and loaded with anti-caries DNA vaccine pGJGLU/VAX. Mice were divided into four groups and immunized through the nasal route with pGJGLU/VAX-loaded BG (Group Ghost+pGJGLU/VAX), pVAX1-loaded BG (Group Ghost+pVAX1), pGJGLU/VAX-Bupivacaine complex (Group pGJGLU/VAX) and pVAX1-Bupivacaine complex (Group pVAX1), respectively. Enzyme-linked immunosorbent assay (ELISA) was used to evaluate the immune responses.

RESULTSELISA results showed that group Ghost+pGJGLU/VAX had significantly higher level of specific anti-GLU SIgA antibody [(0.367 ± 0.086) A/µg] compared with group Ghost+pVAX1 [(0.122 ± 0.077) A/µg], Group pGJGLU/VAX[(0.068 ± 0.068) A/µg] or Group pVAX1[(0.089 ± 0.089) A/µg] (P = 0.028, 0.012 or 0.030, respectively).

CONCLUSIONSSt ghost was developed successfully, which enhanced the efficacy of immune responses induced by anti-caries DNA vaccine pGJGLU/VAX via the nasal route.

Animals ; Antibodies, Antinuclear ; Bacterial Vaccines ; Dental Caries ; prevention & control ; Immunoglobulin A, Secretory ; Mice ; Plasmids ; Salmonella typhimurium ; Vaccines, DNA

Typhoid fever, a serious systemic infection caused by Salmonella enterica serovar Typhi, breaks out in developing countries. However, existing vaccines only induce relatively low protective effects with humoral responses and do not stimulate secondary immune response, especially to young people. The objective of this study is to evaluate the immunogenicity of the vaccine containing virulence capsular polysaccharide (Vi) conjugated with the optimal ratios of non-toxic variant of diphtheria toxin (CRM(197)) in mice. Six-week-old BALB/c female mice were injected intraperitoneally three times at intervals of 14 days and sera were collected on days 0, 14, 28, 42 and 56 post-injection. The efficacy of the vaccine was evaluated by comparing between negative control group injected with PBS and vaccine groups injected with Vi or Vi-CRM(197) conjugate of different ratio. Vi and CRM(197)-specific antibody responses were evaluated using enzyme-linked immunosorbent assay. The result showed that Vi-CRM(197)-1 group revealed the highest and significant Vi-specific IgG immune responses among the other groups and Vi group (p < 0.01). In conclusion, Vi-CRM(197)-1 conjugate vaccine induced the highest humoral immune response in mice and may be used as an effective vaccine to replace the existing typhoid vaccine for infants under 2 years old.
Animals ; Antibody Formation ; Child, Preschool ; Developing Countries ; Diphtheria Toxin ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Immunity, Humoral ; Immunoglobulin G ; Infant ; Mice* ; Salmonella enterica* ; Salmonella typhi* ; Salmonella* ; Typhoid Fever ; Typhoid-Paratyphoid Vaccines ; Vaccines ; Virulence

Salmonella enterica Gallinarum (SG) causes fowl typhoid (FT), a septicemic disease in avian species. We constructed deletion mutants lacking the stress sigma factor RpoS, the nitric oxide (NO)-detoxifying flavohemoglobin Hmp, and the SsrA/SsrB regulator to confirm the functions of these factors in SG. All gene products were fully functional in wild-type (WT) SG whereas mutants harboring single mutations or a combination of rpoS, hmp, and ssrAB mutations showed hypersusceptibility to H2O2, loss of NO metabolism, and absence of Salmonella pathogenicity island (SPI)-2 expression, respectively. A triple-deletion mutant, SGDelta3 (SGDeltarpoSDeltahmpDeltassrAB), was evaluated for attenuated virulence and protection efficacy in two-week-old Lohmann layer chickens. The SGDelta3 mutant did not cause any mortality after inoculation with either 1 x 10(6) or 1 x 10(8) colony-forming units (CFUs) of bacteria. Significantly lower numbers of salmonellae were recovered from the liver and spleen of chickens inoculated with the SGDelta3 mutant compared to chickens inoculated with WT SG. Vaccination with the SGDelta3 mutant conferred complete protection against challenge with virulent SG on the chickens comparable to the group vaccinated with a conventional vaccine strain, SG9R. Overall, these results indicate that SGDelta3 could be a promising candidate for a live Salmonella vaccine against FT.
Administration, Oral ; Animals ; Bacterial Proteins/*genetics/immunology ; *Chickens ; Female ; Poultry Diseases/*immunology/microbiology ; Salmonella Infections, Animal/*immunology/microbiology ; Salmonella Vaccines/administration & dosage/genetics/*immunology ; Salmonella enterica/immunology/*physiology ; Vaccines, Attenuated/administration & dosage/genetics/immunology ; Virulence

The fusion protein (F) gene of Newcastle disease virus was amplified by polymerase chain reaction (PCR) from the recombinant plasmid pVAX1-F, and subcloned into eukaryotic expression vector pmcDNA3. 1+. The F gene was identified by sequencing. The recombinant plasmid was transformed into attenuated Salmonella typhimurium SL7207, and the recombinant was designated as SL7207 (pmcDNA3. 1-F). In vitro and in vivo experiments showed that the plasmid stability of pmcDNA3. 1-F was apparently higher than that of pcDNA3. 1-F in SL7207. In order to compare the immune response induced by these two re combinant bacteria, BALB/c mice were immunized orally with them at the dosage of 2 x 10(9) CFU respectively. Both SL7207(pcDNA3. 1-F) and SL7207(pmcDNA3. 1-F) initiated F-specific serum and mucosal antibodies in immunized mice. Furthermore, 4-day-old SPF chickens were immunized with SL7207(pcDNA3. 1-F) and SL7207(pmcDNA3. 1-F) at the dosage of 5 x 10(9) CFU and boosted two weeks later with the same dosage. Humoral and intestinal mucosal immune responses were observed and their levels were significantly higher than that of negative and positive controls. The result of protective efficacy showed that the chickens immunized with SL7207(pmcDNA3. 1-F) had the protective rate of 70.0%, higher than that of the SL7207 (pcDNA3. 1-F) with 50.0%. In summary, the DNA vaccine delivered by attenuated Salmonella typhimurium has good immunogenicity. A novel mucosal DNA vaccine has been developed and could be useful for controlling the infection and epidemic of Newcastle disease in the poultry.
Animals ; Chickens ; Female ; Immunization ; Mice ; Mice, Inbred BALB C ; Newcastle disease virus ; immunology ; Plasmids ; Salmonella typhimurium ; genetics ; Vaccines, Attenuated ; immunology ; Vaccines, DNA ; immunology ; Viral Vaccines ; immunology

Salmonella enterica serovar Choleraesuis strain C500 is a live, attenuated vaccine that has been used in China for over 40 years to prevent piglet paratyphoid. The objective of this study was to evaluate the potential of attenuated Salmonella enterica serovar Choleraesuis C500 strain with a delta asd mutant as an effective live vaccine vector by the Asd+ balanced-lethal host-vector system. Here, we compared the characteristics of S. enterica serovar Choleraesuis delta asdC500 strain with the parent C500 strain, including phenotype, growth rate, virulence, safety, and expression for heterologous antigen. The mean generation times of delta asdC500 mutant, the vector control delta asdC500 (pYA3493), and the parent avirulent C500 vaccine strain in Luria broth were 30.7, 28.1, and 27.9 min, respectively. The fermentation patterns of theses three strains on different carbohydrates, and the levels of production of H2S, were similar. The O and H antigens of delta asdC500 mutant, delta asdC500 (pYA3493) and delta asdC500 (pYA-F1P2) were 6,7:C:1,5, identical to the parent strain C500. By the method of Reed and Muench, groups of mice were challenged by the intraperitoneal route with different amounts of delta asdC500 (pYA3493) or the parent C500 strain, and the virulence of delta asdC500 (pYA3493) with LD50 of 1.1 x 10(7) CFU was a little lower than C500 with LD50 of 4.4 x 10(6) CFU. All piglets inoculated with delta asdC500 (pYA3493) or C500 survived, and no signs of disease were observed during the entire experimental period. No major differences were found in these two groups. In addition, the recombinant pYA-F1P2 plasmid was very stable in the recombinant delta asdC500 (pYA-F1P2) strain, which expressed secretorily a large amount of the recombinant filamentous hemagglutinin type I domain and pertactin region 2 domain antigen (rF1P2) of Bordetella bronchiseptica. In this study, we have shown that the delta asdC500 mutant had a series of biological characteristics similar to the parent vaccine strain C500. Furthermore, the strain could express secretorily a large amount of heterologous antigen. It is likely that this Salmonella expression and delivery system could be easily adapted to develop multivalent recombinant Salmonella vaccines against infectious agents using the Asd+ balanced-lethal host-vector system.
Amino Acid Oxidoreductases ; genetics ; Animals ; Bacterial Proteins ; genetics ; Gene Deletion ; Genetic Vectors ; Mice ; Mutation ; Salmonella Vaccines ; genetics ; immunology ; Salmonella enterica ; genetics ; immunology ; pathogenicity ; Swine ; Transduction, Genetic ; Vaccines, Attenuated ; genetics ; immunology ; Vaccines, Synthetic ; genetics ; immunology ; Virulence

Interleukin-18 (IL-18) has been known to induce interferon-gamma (IFN-gamma) production and promote Th1 immunity. Although mammalian IL-18 has been characterized in great detail, the properties and application of chicken IL-18 remain largely uninvestigated as of yet. In this study, we evaluated the immunomodulatory properties of Salmonella enterica serovar Typhimurium expressing chicken interleukin-18 (chIL-18) on immune responses induced by avian influenza (AI) and Newcastle disease (ND) vaccines. After oral administration of S. enterica serovar Typhimurium expressing chIL-18, chickens were vaccinated intramuscularly with the recommended dose of either inactivated AI H9N2 vaccine or ND (B1 strain) vaccine. Chickens receiving a primary vaccination were boosted using the same protocol 7 days later. Humoral and cell-mediated immune responses were evaluated in terms of HI antibody titers and proliferation and mRNA expression of IFN-gamma and IL-4 of peripheral blood mononuclear cells (PBMC) in response to specific antigen stimulation. According to our results, oral administration of S. enterica serovar Typhimurium expressing chIL-18 induced enhanced humoral and Th1-biased cell-mediated immunity against AI and ND vaccines, compared to that of chickens received S. enterica serovar Typhimurium harboring empty vector. Therefore, we conclude that our proposed vaccination regimen using inactivated AI and ND viruses along with oral administration of S. enterica serovar Typhimurium expressing chIL-18 may provide a novel approach in protecting chicken from currently circulating AI and ND virus strains.
Administration, Oral ; Animals ; Chickens ; Immunity, Cellular ; Influenza in Birds ; Interferon-gamma ; Interleukin-18 ; Interleukin-4 ; Newcastle Disease ; RNA, Messenger ; Salmonella ; Salmonella enterica ; Vaccination ; Vaccines ; Viruses