In this study, a genetically engineered live attenuated Salmonella Enteritidis (SE) vaccine was evaluated for its ability to protect against Salmonella Typhimurium (ST) infection in chickens. The birds were orally primed with the vaccine on the 1st day of life and given an oral booster at 5 wk of age. Control birds were orally inoculated with phosphate-buffered saline. Both groups of birds were orally challenged with a virulent ST strain at 9 wk of age. Compared with the control chickens, the vaccinated chickens had significantly higher levels of systemic IgG and mucosal IgA against specific ST antigens and a significantly greater lymphoproliferative response to ST antigens. The excretion of ST into the feces was significantly lower in the vaccinated group than in the control group on days 9 and 13 d after challenge. In addition, the vaccinated group had significantly fewer pronounced gross lesions in the liver and spleen and lower bacterial counts in the internal organs than the control group after challenge. These data indicate that genetically engineered live attenuated SE may induce humoral and cellular immune responses against ST antigens and may confer protection against virulent ST challenge.

A temporal study was carried out to determine Salmonella prevalence, trends, major serovars, and their clusters from environmental samples, in poultry breeder flocks in Ontario between January 1998 and December 2008. Surveillance data were obtained from the Ontario Hatchery and Supply Flock Policy. Logistic regression with a random effect for flock was used to identify factors [poultry type, year (trend) and season] associated with the prevalence of Salmonella. A cluster detection test was used to identify clusters of common serovars. The period prevalence of Salmonella was 47.4% in broiler-breeder, 25.7% in layer-breeder, and 19.6% in turkey-breeder flocks. The overall trend in the prevalence of Salmonella was decreasing for all breeder types, due primarily to decreasing trends of Salmonella Heidelberg. The seasonal effects varied by year with the highest probability of Salmonella occurring in different seasons. The 4 most common serovars identified were Salmonella Heidelberg, Kentucky, Hadar, and Typhimurium in broiler-breeders; Salmonella Heidelberg, Brandenburg, Thompson, and Typhimurium in layer-breeders; and Salmonella Heidelberg, Saintpaul, Brandenburg, and Muenster in turkey-breeders. Salmonella Enteritidis was infrequently isolated in all poultry breeder types. Temporal clusters of different serovars were identified in all poultry breeder types. Clusters of Salmonella Heidelberg, Typhimurium, and Hadar from environmental samples from breeder flocks were detected during a similar period to clusters from hatchery fluff samples from the same population. Therefore, interventions at the breeder flock-level might help to reduce transmission of Salmonella from breeder flocks to hatcheries and possibly, to lower levels of the poultry production chain.

The innate immune system is a critical component directing the overall response of the immune system early in the inflammatory process. Evaluation of the innate immune system could offer a screening method for the selection of breeding stock from commercial chicken operations to improve flock health and prevent the loss of genes crucial to disease resistance. Three commercial broiler chicken lines (designated Lines A, B and C) were profiled for efficiency of their innate immunologic response. Oxidative burst and bactericidal functions of heterophils and monocytes, as well as heterophil degranulation, were analyzed. The birds were tested 1, 4, 8 and 15 days post-hatch. Individual lines differed in their ability to perform innate immunological responses during the first 15 days post-hatch. Although bactericidal capabilities were similar, oxidative burst responses by monocytes were low in comparison to that generated by heterophils. The fact that monocytes are not particularly adept at producing an oxidative burst at this age suggests that this is not a major avenue of innate defense by monocytes. Heterophil oxidative burst response was stronger in Line C than Line A during the first four days post-hatch. Line B showed no difference from Line C in heterophil oxidative burst response at 1 d, but produced a stronger response than Line C on 4 and 8 d post-hatch. Degranulation by heterophils showed significant differences in responses of Lines A and C depending on the day post-hatch, and stronger response in Line C vs Line B in the first four days post hatch. The first week post-hatch is an important time as chicks are particularly susceptible to infection as neonates. Mortality data of the commercial lines indicates that Line A is the most susceptible to demise, followed by Line C and then Line B. These results suggest that oxidative burst production efficiency is an important defensive function to monitor for immunoprofiling.

Virulence of three Canadian poultry strains of Salmonella enteritidis, namely phagetypes (PT) 4, 8 and 13, and one Salmonella heidelberg strain was assessed in orally and intraperitoneally inoculated one-day old chickens and compared to the virulence of a human S. enteritidis PT 4 strain from the United Kingdom (UK). The two PT 4 strains were also compared in orally inoculated adult laying hens. In addition, orally inoculated Balb/c mice were used to evaluate virulence of the above strains and two strains of Salmonella typhimurium containing different plasmids. In orally inoculated one-day old chickens, the UK S. enteritidis PT 4 strain was more virulent than the Canadian PT 4 strain. The UK PT 4 strain was also more virulent and invasive in adult laying hens than the Canadian PT 4 strain. The S. enteritidis PT 8 strain and one S. typhimurium strain isolated from a chicken hatchery were the most virulent for orally inoculated Balb/c mice. This strain of S. typhimurium contained the 60 megadalton plasmid associated with virulence for Balb/c mice which was not present in the S. typhimurium strain isolated from a pig with septicemic disease.

This study compared a new live attenuated Salmonella Enteritidis vaccine candidate secreting Escherichia coli heat-labile enterotoxin B subunit (SE-LTB) with a commercial Salmonella Enteritidis (SE) vaccine for efficacy of protection against SE infection in laying hens. Chickens were divided into 3 groups of 20 each. Group A chickens were inoculated orally with phosphate-buffered saline and served as controls, group B chickens were inoculated orally with the vaccine candidate, and group C chickens were inoculated intramuscularly with a commercial vaccine, the primary inoculation in groups B and C being at 10 wk of age and the booster at 16 wk. Groups B and C showed significantly higher titers of plasma immunoglobulin G, intestinal secretory immunoglobulin A, and egg yolk immunoglobulin Y antibodies compared with the control group, and both vaccinated groups showed a significantly elevated cellular immune response. After virulent challenge, group B had significantly lower production of thin-shelled and/or malformed eggs and a significantly lower rate of SE contamination of eggs compared with the control group. Furthermore, the challenge strain was detected significantly less in all of the examined organs of group B compared with the control group. Group C had lower gross lesion scores only in the spleen and had lower bacterial counts only in the spleen, ceca, and ovary. These findings indicate that vaccination with the SE-LTB vaccine candidate can efficiently reduce internal egg and internal organ contamination by Salmonella and has advantages over the commercial vaccine.

The humoral and cell-mediated immune (CMI) response to 2 commercial killed Salmonella Enteritidis (SE) vaccines (Layermune and MBL SE4C) was evaluated in laying hens. Layers were distributed in 2 experimental groups. The first received a single immunization at 16 wk of age, while the second experimental group was immunized at 12 wk of age and again at 18 wk of age. Serum immunoglobulin (Ig)G antibodies were measured using a commercial SE ELISA kit and showed persistent levels from 3 to 32 and 34 wk post-vaccination. The vaccination protocol using 2 immunizations showed a higher seroconversion level than the single vaccination. However, our results for bacterial intracellular survival indicated that IgG titers were not linked with bacterial killing. Local IgA production was measured in the intestines and oviducts with an in-house SE whole cell antigen ELISA. Only the MBL SE4C vaccine elicited IgA antibody production when tested on intestine and oviduct mucosal secretions, 3-weeks post-vaccination in both immunization protocol groups. To evaluate the CMI response, the splenic T-cells and B-cells populations were analyzed using flow cytometry. The CD3/B-cell ratio decreased 3 wk after the second immunization in the twice vaccinated Layermune group due to an increase in B-cells.

Liposomes and immunostimulating complexes (ISCOM) are adjuvants that have been known to potentiate the immune response to membrane proteins. Adjuvanted outer membrane proteins (OMP) from Salmonella enteritidis were evaluated for their protective efficacy against S enteritidis infection in turkeys. The adjuvanted vaccines prepared for evaluation were: positive or negatively charged liposomes, lipid-conjugated ISCOM, and mineral oil vaccines. These preparations were compared with that of a whole cell bacterin and protein alone. After vaccination, turkeys were challenge-exposed with a nalidixic acid-resistant strain of S enteritidis. They were monitored for clinical signs of disease, antibody response, bacterial shedding pattern, and clearance of the challenge S enteritidis from internal organs. Results indicated a significantly (P < 0.05) higher antibody response to the positively charged liposomal OMP vaccine, compared with the whole cell bacterin. The antibody response to positively charged liposomal OMP vaccine was greater when a booster dose of this preparation was given. Shedding of S enteritidis was decreased in all vaccinated and challenge-exposed turkeys (P < 0.001). The tissues from a high percentage (90 to 100%) of birds that received a booster vaccination of the liposomal (+ or -) and ISCOM vaccine were culture-negative for S enteritidis.

An ELISA was developed and tested for its ability to detect antibodies against Salmonella enteritidis in chickens. Various features of the ELISA were evaluated and optimized. The outer membrane protein antigens selected by use of the protein immunoblotting method made the assay specific and sensitive. The assay was evaluated in chickens experimentally infected with S enteritidis. Blood samples collected at weekly intervals after experimental infection with S enteritidis were analyzed by ELISA. Results of the ELISA were compared with those of conventional serum plate and microagglutination tests. The ELISA was more sensitive and specific in the detection of S enteritidis infection than the other 2 conventional tests.

The purpose of this study was to evaluate the effectiveness of an aromatic-dependent mutant of Salmonella typhimurium as a parenteral vaccine for prevention of fecal shedding of Salmonella spp. Pigs and chickens were vaccinated IM, with 1 X 10(9) and 1 X 10(8) organisms, respectively, followed by a second identical vaccination 2 weeks later. Salmonella organisms were not detected by analysis of fecal or cloacal swab specimens from any animal after vaccination. Deleterious side effects were not noticed after vaccination. Pigs were challenge-inoculated PO with 1 X 10(12) virulent S typhimurium 1 week after the second vaccination. Chickens were challenge-inoculated PO with 3 X 10(8) organisms of either S enteritidis or the virulent parent strain of S typhimurium 3 weeks after the second vaccination. Vaccinated pigs shed Salmonella spp significantly less frequently than did nonvaccinated pigs. Vaccinated chickens challenge-inoculated with either S enteritidis or S typhimurium also shed Salmonella less frequently than the corresponding nonvaccinated control birds; however, the difference was not significant.

The aim of this study was to evaluate the antimicrobial activity and determine the minimum bactericidal concentration (MBC) of the essential oils derived from Origanum vulgare (oregano), Melaleuca alternifolia (tea tree), Cinnamomum cassia (cassia), and Thymus vulgaris (white thyme) against Salmonella Typhimurium, Salmonella Enteritidis, Escherichia coli, Staphylococcus aureus and Enterococcus faecalis. The study also investigated the ability of these different bacterial strains to develop adaptation after repetitive exposure to sub-lethal concentrations of these essential oils. The MBC of the essential oils studied was determined by disc diffusion and broth dilution methods. All essential oils showed antimicrobial effect against all bacterial strains. In general, the development of adaptation varied according to the bacterial strain and the essential oil (tea tree > white thyme > oregano). Therefore, it is important to use essential oils at efficient bactericidal doses in animal feed, food, and sanitizers, since bacteria can rapidly develop adaptation when exposed to sub-lethal concentrations of these oils.