Efficacy of a live avirulent Salmonella typhimurium vaccine in preventing colonization and invasion of laying hens by Salmonella typhimurium and Salmonella enteritidis

An avirulent live delta cya delta crp Salmonella typhimurium strain chi 3985 that precludes colonization and invasion of chickens by homologous and heterologous Salmonella serotypes was evaluated for its long-term protection efficacy. Chickens vaccinated orally at 2 and 4 wk of age were assessed for protection against oral challenge with wild-type S. typhimurium and Salmonella enteritidis strains at 3, 6, 9, and 12 mo of age. A comparison of Salmonella isolation from vaccinated and nonvaccinated layers after challenge with S. typhimurium or S. enteritidis showed that delta cya delta crp S. typhimurium chi 3985 induced excellent protection against intestinal, visceral, reproductive tract, and egg colonization, invasion, and/or contamination by Salmonella. The duration of protection lasted for 11 mo after vaccination, at which time the experiment was terminated. S. enteritidis and S. typhimurium were isolated from the yolk, albumen, and shells of eggs laid by nonvaccinated chickens challenged with Salmonella. S. typhimurium caused pathological lesions in nonvaccinated chickens, whereas vaccinated and nonvaccinated chickens challenged with S. enteritidis showed no pathological lesion in the visceral and reproductive organs. Vaccination with chi 3985 prevented transmission of S. typhimurium or S. enteritidis into eggs laid by vaccinated layers with no effect on egg production. To our knowledge, this is the first publication confirming that vaccination with live avirulent Salmonella can induce long-term protection against Salmonella infection in layers.     

Evaluation of bacterins containing three predominant phage types of Salmonella enteritidis for prevention of infection in egg-laying chickens

Six Salmonella enteritidis bacterin formulations differing in adjuvant content and whole-cell inactivation procedures were evaluated in egg-laying chickens. Chickens given S enteritidis bacterins containing modified Freund's incomplete adjuvant had greater humoral immune responses to S enteritidis than did birds given other bacterin formulations (P < 0.05). Better protection against infection by S enteritidis phage types 8, 13a, and 23 was obtained in birds vaccinated with bacterin 5. Bacterin 5 contained S enteritidis cells inactivated by 20% acetone and modified Freund's incomplete adjuvant.     

Antibodies to Klebsiella pneumoniae lipopolysaccharide in patients with ankylosing spondylitis

The role of microbial lipopolysaccharides (LPS) in the aetiopathogenesis of ankylosing spondylitis (AS) is a matter of continuing debate. In this study, class-specific IgG, IgA and IgM antibodies against Klebsiella pneumoniae, Escherichia coli, Salmonella typhimurium and Salmonella enteritidis LPS were measured by enzyme-linked immunosorbent assay (ELISA) in 100 AS patients, 50 rheumatoid arthritis (RA) patients and 50 healthy control subjects. The AS patients had significantly elevated levels of IgG and IgA antibodies against K. pneumoniae LPS (P < 0.001) and IgA antibodies against E. coli LPS (P < 0.05) compared to healthy controls. There were no significant elevations of antibody levels against S. typhimurium and S. enteritidis in the three study groups. In addition, there was a correlation between IgG and IgA anti-K. pneumoniae LPS antibody levels and the acute-phase reactant C-reactive protein (P < 0.001).     

Analysis of splenic and thymic lymphocyte subpopulations in chickens infected with Salmonella enteritidis

Lymphocytes expressing CD3, CD4, CD8, pan lymphocyte, IgA, IgG and IgM cell surface antigens were assessed by in the spleen and thymus of chickens following infection with Salmonella enteritidis using flow cytometric analysis. At 6 days post primary infection and 2 days post secondary infection with S. enteritidis, the percentages of IgA+ and IgM+ lymphocytes in the spleen were significantly increased (P < 0.05). At 2 days post secondary infection with S. enteritidis, the percentage of CD4+ T lymphocyte in the spleen and CD8+ T lymphocyte percentage in the thymus were significantly increased (P < 0.05). These results indicate that S. enteritidis infection induces changes in the spleen and thymus that reflect the dynamics of the host protective immune response.     

Salmonella enteritidis colonization in turkey poults

An experimental trial on the colonization of two Salmonella enteritidis strains (phage types 4 and 8) originating from meat turkey flocks, were carried out. Three-day old poults were inoculated orally with approximately 10(6) cfu/bird (one group with PT8 and the other with PT4). clinical signs were not observed in any of the groups. The total reisolation rates from cloacal swabs in birds inoculated with PT4 was higher than from birds inoculated with PT8. The inoculated strains could also be detected in contact birds. Examination of internal organs 21 days p.i. revealed higher isolation rates in the group infected with PT4 than with PT8. The results indicate that both phage types of S. enteritidis were able to colonize the intestinal tract and the internal organs of turkey poults. However, the colonization and duration of shedding differed between the two phage types. This may be due to the fact that PT8 strain used was free from the S. e. virulence plasmid 37 MDa.     

Yeast treatment to reduce Salmonella and Campylobacter populations associated with broiler chickens subjected to transport stress

The prevalence of Campylobacter and Salmonella on retail poultry carcasses remains a significant public health concern. The stresses associated with transporting poultry prior to slaughter have been shown to increase pathogen populations both in the intestinal tract and on the carcass exterior. The yeast, Saccharomyces boulardii, was evaluated for its ability to reduce populations of Salmonella and Campylobacter in broiler chickens subjected to transport stress. Chicks, inoculated with individual strains of Salmonella and Campylobacter were held for 6 wk and then divided into two groups with half of the chickens receiving 10% dried yeast in the feed for 60 h. The birds were then caged and transported to simulate commercial conditions. After euthanatizing the birds, the ceca were aseptically removed and analyzed for Salmonella and Campylobacter. With no yeast treatment, transport stress caused the Salmonella colonization frequency to increase more than fivefold, from 3.3 to 16.7%. Yeast treatment significantly reduced the frequency of Salmonella colonization to lower than prestress levels, as no Salmonella were recovered from the ceca of these birds (P < 0.05). Similar results were obtained from birds challenged with a mixture of Salmonella and Campylobacter strains. Before transport, 53.3% of these chickens were positive for Salmonella. Transport stress increased the colonization rate to 67.5% in control birds, whereas the colonization of yeast-treated chickens decreased to 40% (P < 0.05). Frequency of Campylobacter isolation from the ceca was not affected by treatment, but Campylobacter populations present in the ceca were significantly reduced in the mixed strain trial (P < 0.05).     

Oral immunization with attenuated Salmonella expressing human sperm antigen induces antibodies in serum and the reproductive tract

Induction of immune responses in the reproductive tract will be crucial for a functional gamete antigen-based antifertility vaccine. Here we describe the construction and development of an avirulent Salmonella as an oral vaccine delivery vector to elicit sperm-specific immune responses in reproductive tract secretions. A cDNA sequence encoding the human sperm antigen SP10 was cloned on an asd+vector and expressed to a high level in an avirulent delta cya, delta crp, and delta asd vaccine strain of Salmonella typhimurium. Oral immunization of female BALB/c mice with this recombinant Salmonella elicited high-titer anti-SP10 IgG antibodies in serum and IgA antibodies in vaginal secretions. Anti-SP10 antibody titers could be increased by secondary and tertiary oral administrations of the recombinant Salmonella. Induction of sperm-specific antibodies in the reproductive tract following oral administration of a recombinant Salmonella could lead to the development of a simple, safe, efficient, and easy-to-use antifertility vaccine.     

Effect of induced molting on the severity of intestinal lesions caused by Salmonella enteritidis infection in chickens

A study was conducted to describe the intestinal lesions caused by Salmonella enteritidis infection in 20-, 40-, and 74-week-old white leghorn chickens that were undergoing a feed deprivation-induced molt. The chickens were infected on the fourth day after feed was removed. At 4 days postinfection (8 days of feed deprivation), cecal and cecal tonsil inflammation was significantly greater in molted infected chickens than in unmolted infected chickens. The cecal lamina propria and epithelium of molted infected chickens contained heterophilic infiltrates, and there were heterophils and sloughed epithelial cells in cecal lumina. Colonic inflammation, consisting of heterophils infiltrating lamina propria and epithelium, occurred more often in molted infected chickens than in unmolted infected chickens. Immunoperoxidase staining of intestinal sections from 20- and 40-week-old chickens revealed S. enteritidis antigen in the lamina propria of cecum, cecal tonsil, and occasionally the colon of molted infected chickens. The character of the S. enteritidis-induced intestinal lesions associated with molting was similar for different ages of birds.     

Effects of gastrin on the histamine-secretory and proliferative activity of cultured carcinoid cells derived from the stomach of the rodent Mastomys natalensis

In order to estimate the distribution of Salmonella including Salmonella enteritidis (SE) and SE-antibodies in commercial layer hen flocks in Hokkaido, the northern prefecture of Japan, a survey of spent layer hens was performed, from August 1996 to January 1997. From the three spent hen processing plants, samples of intestines and sera were collected from 740 birds presented for slaughter from 37 flocks of 22 layer hen farms. Intestines from each birds were cultured for Salmonella including Salmonella enteritidis. Serum from each bird was examined for SE-antibody with the enzyme-linked immunosorbent assay (ELISA). Salmonella (any serotype) and Salmonella enteritidis were isolated from 50 (6.8%) and three (0.4%) of 740 birds, respectively, and SE-antibody positive values were recorded from seven birds (0.9%). SE-antibody positive birds did not always indicate isolation of Salmonella enteritidis, however SE-antibody positive hens were demonstrated only from Salmonella enteritidis positive flocks. Salmonellae were isolated from the birds of ten layer hen farms, all of these hens were raised in houses without windows and with automatic feeders. No isolations of salmonella were made from birds raised in houses with windows. From the windowless houses, Salmonellae were isolated from 46 (21.8%) of 260 birds in houses with four to six cages piled up vertically, and from six (2.5%) of 240 samples from the houses with four to five cages piled in a slanting manner.     

Protection of mice against gastric colonization by Helicobacter pylori by single oral dose immunization with attenuated Salmonella typhimurium producing urease subunits A and B

Helicobacter pylori is a Gram-negative bacterial pathogen associated with gastritis, peptic ulceration, and gastric carcinoma. The bacteria express a strong urease activity which is known to be essential for colonization of gnotobiotic pigs and nude mice. UreA and UreB, two structural subunits of the active enzyme, were expressed in the attenuated Salmonella typhimurium live vaccine SL3261 strain. Evaluation of protection against H. pylori was performed in Balb/c mice by oral immunization with a single dose of the vaccine strain. Five weeks after immunization, mice were challenged orally three times with a mouse-adapted H. pylori wild type strain and, six weeks later, mice were sacrificed to determine H. pylori infection by detection of urease activity from the antral region of the mouse stomachs. In several independent experiments, we observed 100% infection with H. pylori in the non-immunized mice and no infection (100% protection) in the mice immunized with S. typhimurium expressing recombinant UreA and UreB. Specific humoral and mucosal antibody responses against UreA and UreB were observed in mice immunized as indicated by western blots and ELISA assays. These data shows that oral immunization of mice with urease subunits delivered by an attenuated Salmonella strain induced a specific immune response and protected mice against H. pylori colonization. Single oral dose immunization with UreA and UreB delivered by a live Salmonella vaccine vector appears to be an attractive candidate for human vaccination against H. pylori infection. In addition, this model will aid to elucidate the effective protection mechanisms against H. pylori in the gastric mucosa.     

Serological and bacteriological observations on experimental infection with Salmonella hadar in chickens

Over the past 3 years the frequency of Salmonella hadar infections has increased in Belgium in both poultry and humans. Therefore, the course of infection with S. hadar in poultry was investigated. One day-old and 4 week-old specific pathogen-free chickens were orally infected with one of two S. hadar strains, SH1 or SH2. Mortality was 6% (SH1) and 17% (SH2) in birds infected at 1 day-old. Chickens infected at 1 day-old with SH2 showed a mild diarrhoea. The S. hadar faecal excretion in birds infected at 1 day-old remained high throughout the experiment until 12 weeks post-inoculation (pi). Faecal excretion was lower in older birds. Antibodies to S. hadar were observed from 2 weeks pi (SH2, infected at 1 day-old) or 4 weeks pi (SH1, both groups; SH2, chickens infected at 4 weeks of age). The percentage of chickens with antibodies was higher after infection at 1 day-old than after infection at 4 weeks of age. In a second experiment 1 day-old chicks were infected with SH1 and autopsied at regular intervals until 42 days pi. SH1 was isolated from the caeca from 3 h pi onwards and from the liver and spleen from 18 h until 14 days pi. Serous typhlitis and omphalitis were the main lesions. The number of macrophages in the lamina propria of the caecal tonsils was slightly increased from 18 h until 2 weeks pi. In the liver, inflammation was observed in the portal triads and in the sinusoids. This study indicates that infections with S. hadar lead to intense colonisation of the gut and extensive faecal shedding. It may also cause invasive infections in 1 day-old chickens.     

Serologic responses of young colostrum fed dairy calves to antigens of Pasteurella haemolytica A1

The potential and limitations of early calfhood vaccination to induce active immunity to Pasteurella haemolytica A1 in conventional colostrum fed calves were investigated. Holstein dairy calves (n = 29) were vaccinated at 2 and 4 weeks of age, or at 6 and 8 weeks of age with a commercial culture supernatant vaccine (Presponse, Langford Inc., Guelph, Ont., Canada), or remained unvaccinated as controls. Serum antibody titres were measured using an indirect bacterial agglutination assay, a leukotoxin neutralization assay, and enzyme immunoassays for antibodies of the IgM, IgG1, and IgG2 isotypes binding purified capsular polysaccharide of P. haemolytica A1. Seroconversion (fourfold or greater increase in serum antibody titre) rates were compared using Fisher's exact test. The effects of passive antibody titres and age on response to vaccination were assessed by linear modelling. Vaccination at 2 and 4 weeks of age was associated with 40%, and 0% of calves seroconverting on the basis of agglutinating antibody titres, and leukotoxin neutralizing titres respectively, and 50%, 0%, and 0% seroconverting on the basis of IgM, IgG1 and IgG2 antibodies to capsular polysaccharide, respectively. Agglutinating antibody responses were not related to prevaccination antibody titres, or to age at vaccination. Higher responses (p = 0.08) to leukotoxin were observed in older calves (after taking differences in prevaccination titres into account). Statistical analyses of responses to capsular polysaccharide among calves with comparable prevaccination IgG1 antibody titres revealed significantly higher IgM, IgG1 and IgG2 responses in older calves. Rising titres of IgM antibodies in nonvaccinated calves after 5 weeks of age suggest natural exposure to P. haemolytica A1 or antigens which result in serologic cross-reactions as a means of priming immune responses.     

Identification and sequence analysis of lpfABCDE, a putative fimbrial operon of Salmonella typhimurium

A chromosomal region present in Salmonella typhimurium but absent from related species was identified by hybridization. A DNA probe originating from 78 min on the S. typhimurium chromosome hybridized with DNA from Salmonella enteritidis, Salmonella heidelberg, and Salmonella dublin but not with DNA from Salmonella typhi, Salmonella arizonae, Escherichia coli, and Shigella serotypes. Cloning and sequence analysis revealed that the corresponding region of the S. typhimurium chromosome encodes a fimbrial operon. Long fimbriae inserted at the poles of the bacterium were observed by electron microscopy when this fimbrial operon was introduced into a nonpiliated E. coli strain. The genes encoding these fimbriae were therefore termed lpfABCDE, for long polar fimbriae. Genetically, the lpf operon was found to be most closely related to the fim operon of S. typhimurium, both in gene order and in conservation of the deduced amino acid sequences.     

Evaluation of the chicken crop as a source of Salmonella contamination for broiler carcasses

Much previously published research has focused on the role of cecal and intestinal Salmonella contamination of poultry carcasses within commercial processing plants. Presently, we have evaluated the persistence of experimentally inoculated Salmonella enteritidis in the crops and ceca of commercial broiler chickens during the last week of growth (Weeks 6 to 7) and the presence of crop and cecal Salmonella in 7-wk-old broilers in a commercial processing plant. When broilers were inoculated with 1 x 10(6) cfu S. enteritidis at 6 wk of age by oral gavage, the incidence of crop and cecal contamination was equivalent 2d after challenge (30%), with only 1 of 29 crops contaminated and 0 of 29 ceca contaminated at 7 d following challenge. When broilers were inoculated with 1 x 10(8) cfu S. enteritidis at 6 wk of age by oral gavage, 2 d after challenge the crops and ceca were observed to be 57 and 67% positive for S. enteritidis, respectively. Seven days after inoculation with 1 x 10(8) S. enteritidis, the crops and ceca were 37 and 57% positive, respectively, for the challenge organism. At a commercial broiler processing plant, 286 of 550 crops from three flocks were Salmonella-positive, whereas only 73 of 500 ceca from these flocks were contaminated. Furthermore, data from this plant indicated that the crops were far more likely to rupture than ceca (86-fold) during processing, increasing the possibility of carcass contamination with Salmonella derived from crop contents. The results of these studies suggest that the crop may serve as a source of carcass contamination with Salmonella within some processing plants.     

Mice as carriers of Salmonella enteritidis on persistently infected poultry units

Evidence of the possible role of wild mice in the epidemiology of Salmonella enteritidis infection on broiler breeder and layer breeder units was obtained by a bacteriological examination of local mice. Persistent S enteritidis infection in birds on these units had been a problem and a high proportion of the mice were found to carry salmonella. S enteritidis was isolated from the liver and the intestine of most of the mice, indicating a systemic infection. Three-week-old chicks were infected by contact with droppings from mice which had been infected experimentally with S enteritidis two and five months previously. Wild mice infected artificially or naturally excreted S enteritidis intermittently, with up to 10(4) organisms in some individual droppings. A naturally infected mouse which died after intermittently excreting small numbers of S enteritidis in its droppings for 19 weeks had 10(4) organisms/g of liver and 10(3)/g of macerated intestine and contents. S enteritidis was also found in fetal tissue in a naturally infected mouse suggesting the possibility that the organism might be transmitted vertically.     

Experimental coryza in broiler chickens. I. Effects of vaccination with Haemophilus gallinarum bacterin and its components on weight gains and resistance to infection

Effects of a Haemophilus gallinarum bacterin and its components were studied in young broiler chickens. When the bacterin was administered subcutaneously in the dorsal neck region at 2 weeks of age, no significant differences in weight gains of vaccinated and control birds were detected at eight weeks of age. In four groups vaccinated at 1, 2, 3, or 4 weeks of age and challenged with virulent haemophilus organisms 3 weeks later, the incidence of clinical signs was 30% in the vaccinates and 60% in controls. The bacterin was equally protective at the four ages of administration. Caseous plugs were found at the vaccination sites in all birds which had received adjuvant either alone or in the complete bacterin. Signs of depression lasting about 24 hours were observed in the youngest birds injected with the bacterin, but overall weight gains were normal.     

Resistance against Salmonella enteritidis cecal colonization in Leghorn chicks by vent lip application of cecal bacteria culture

Leghorn chicks were treated with cultures of cecal bacteria from adult chickens by crop gavage, upper body spray, or vent lip application on the day of hatch. The chicks were challenged orally with 10(4) Salmonella enteritidis (SE) at 3 d of age and evaluated for SE cecal colonization at 10 d of age. The concentration of volatile fatty acids (VFA) in the cecal contents was determined on the day after culture treatment and at 10 d of age. Compared with controls, SE colonization was significantly decreased in each of the treatment groups. Vent lip application of a single .05-mL drop of cecal bacteria culture resulted in resistance against SE challenge comparable to crop gavage or spray treatment with .5 mL of culture. Resistance to SE challenge was directly associated with the concentrations of total VFA and propionic acid in the cecal contents of the treated chicks on the day after culture treatment. The results indicated that cecal bacteria from adult chickens that increase SE colonization resistance may rapidly become established in the ceca of newly hatched chicks following contact with the vent lips.     

Protection of chickens with or without maternal antibodies against both Marek's and Newcastle diseases by one-time vaccination with recombinant vaccine of Marek's disease virus type 1

We constructed a stable recombinant Marek's disease virus type 1 (rMDV1) expressing the fusion protein (F) of Newcastle disease virus (NDV) by inserting the coding sequence within the US10 gene of MDV1 by homologous recombination and designated this as rMDV1-US10L(F). The NDV-F protein was significantly expressed under control of the SV40 late promoter in cultured cells infected with the rMDV1. To examine the protective efficacy of the rMDV1, specific pathogen-free (SPF) chickens were vaccinated with rMDV1 at one-day-old. Almost all birds (> 95%) were protected from NDV challenge via intramuscular, ocular, intranasal and intratrachial routes at 4 weeks after vaccination. The rMDV1 showed 100% protection against virulent MDV1 challenge in SPF chickens. Antibody responses against NDV-F and MDV1 antigens were observed at least up to 11 weeks after immunization. When the sera from chickens vaccinated with the rMDV1 were examined for the presence of anti-NDV-F antibody on the day of NDV challenge, the vaccinated bird group which did not survive from NDV challenge were found to show lower antibody titers than the surviving group. The rMDV1 also provided sufficient protection against NDV and MDV1 challenges in commercial chickens with maternal antibodies against NDV-F and MDV1 antigens.     

Protective immune response of chickens against Newcastle disease, induced by the intranasal vaccination with inactivated virus

Intranasal vaccination of chickens with inactivated Newcastle disease virus (NDV) induced both local and systemic antibody responses, resulting in protection against intranasal challenge with a lethal dose of a virulent NDV strain. The immune response was enhanced by the use of cholera toxin B subunit (CTB) as an adjuvant and only small amounts of the challenge virus were recovered from the birds vaccinated together with CTB. On the other hand, subcutaneous vaccination with the same antigen induced only a serum antibody response in chickens, allowing the challenge virus to replicate in the sinus. The present results indicate that secretory antibodies induced on the respiratory mucosal surface by intranasal vaccination with inactivated NDV protected chickens from lethal infection by inhibiting virus replication at the portal of entry for the virus.     

Salmonella prevalence in crops of Ontario and Quebec broiler chickens at slaughter

Swabs of crop contents of 635 broiler chickens were obtained from 9 Ontario and 12 Quebec processing plants and cultured for Salmonella to determine prevalence in broiler crops. Serotypes of positive cultures were determined to evaluate the serotype profile. The overall prevalence of contamination was low (4.3%). Prevalence was higher in broilers sampled in Quebec (5.8%) than in those sampled in Ontario (2.2%). In Quebec, there were differences in prevalence among the groups of broilers sampled at the various plants. These differences were believed to be attributable to differences in Salmonella prevalence among groups of flocks delivered to the plants due to the limited exposure of the chickens to the plant. The serotype profile of Salmonella isolated from the crops of broilers in this study was similar in several respects to profiles obtained from other surveys of Canadian broiler flocks using either environmental samples or cloacal swabs. Similarities included: predominance of Salmonella hadar and Salmonella heidelberg; several other common serotypes at a low prevalence; little Salmonella enteritidis isolated in other studies, and no S. enteritidis isolated in this study. Results of this field survey of Salmonella in crops of broilers are similar to those of Canadian studies of other internal and environmental sites of broilers. The similarity indicates that monitoring of Salmonella environments of flocks of live broiler chickens should define profiles of Salmonella contamination of the carcasses and would also aid in determination of Salmonella contamination status of broiler flocks. Such information would assist efforts to reduce Salmonella contamination in broiler chickens.