Aromatic-dependent Salmonella typhimurium as modified live vaccines for calves

Strains of Salmonella sp with complete nonreverting aromatic biosynthesis (aro) defects are expected to be nonvirulent, in respect to invasive infection, because they need the aromatic metabolites paraaminobenzoate (for making folate) and dihydroxybenzoate (for making enterochelin) which are not available in host tissues. Derivatives with transposon-generated complete nonreverting aro-defects were prepared from 3 mouse-virulent strains of S typhimurium, namely, FIRN, WRAY, and UCD. The latter 2 parent strains originally were isolated from calves and are known to be calf-virulent. The resultant aromatic-dependent (aro-) strains were used to vaccinate 27 calves (2 to 3 weeks old), usually giving 2 doses by the IM route (10(9) bacteria) or orally (1.5 X 10(11)). Vaccination did not cause severe ill effects in any calf. Thus aro- defects cause loss of virulence for calves, as previously shown for mice. Vaccinated and control calves were challenge exposed, usually at 5 weeks of age, by feeding 1.5 X 10(11) cells of 1 of 2 calf-virulent S typhimurium strains, either UCD 108-11 or SL1323. Of the 16 challenge-exposed control calves, all became anorectic and depressed (CNS), and 15 had diarrhea. Fourteen of the 16 died; all tested tissues were bacteriologically culture-positive for Salmonella at necropsy. Vaccination with the live UCD aro- vaccine strain, SL1479 by either of 2 schedules (IM or orally) appeared effective.(ABSTRACT TRUNCATED AT 250 WORDS)     

Duration of immunity induced in chickens by an attenuated live Salmonella enteritidis vaccine and an inactivated Salmonella enteritidis/typhimurium vaccine

The aim of this study was to examine the duration of immunity of different vaccination schemes using the S. enteritidis live vaccine Gallivac Se and the S. enteritidis-S. typhimurium inactivated vaccine Gallimune Se+St. Three groups of Lohman Brown chickens were used. Group one was vaccinated three times orally with Gallivac Se at weeks one, seven and 13 of age. Group two was vaccinated twice orally with Gallivac Se in weeks one and seven and once i.m. with Gallimune Se+St in week 14 of age. A third group was not vaccinated and served as the control group. Eight randomly selected chickens from each of the three groups were challenged with a nalidixic acid resistant S. enteritidis PT4 strain in weeks 24, 51 and 71 of age and the same number of animals were challenged with a S. typhimurium DT 104 strain in weeks 26, 54 and 73 (75) of age.The chickens were euthanised seven days post challenge and the number of challenge strain organisms (log10 cfu) in the liver and on caecal mucosa was determined.The quantitative investigation of the challenge strain in the liver and caecal mucosa revealed a statistically significant (p < 0.05) lower challenge strain burden in the vaccinated groups compared with the non-vaccinated control group up to week 71 (73) of age. The protective effects were demonstrated for both challenge strains.     

Galactose epimeraseless mutants of Salmonella typhimurium as live vaccines for calves.

The purpose of the study was to evaluate the safety and efficacy of a galactose epimeraseless mutant of Salmonella typhimurium administered as an oral vaccine to one week old calves and to investigate properties of galactose epimeraseless mutants which affect their virulence and immunogenicity. The galactose epimeraseless mutant S. typhimurium strain G30D caused diarrhea and fever in three calves to which it was administered orally at a dose of 10(10) organisms; all three calves died following challenge with virulent S. typhimurium ten days postvaccination. Mild illness developed in four calves vaccinated with a dose of 9 X 10(6) organisms and one of these calves survived challenge. Three unvaccinated calves died following challenge. The vaccine organism persisted in tissues and was shed for a prolonged period by calves which received 10(10) organisms. Studies of characteristics of galactose epimeraseless mutants of S. typhimurium showed that, in the presence of galactose, there is selection for secondary mutants which are galactose resistant. The studies indicate that galactose epimeraseless mutants of S. typhimurium are not good candidate live vaccine organisms for use in calves.     

Immunization of calves with live and inactivated whole-cell vaccines against Salmonella typhimurium infection

Eight calves were immunized with live auxotrophic Salmonella typhimurium mutants (aro -SL 1479, gal E 3821) and twelve calves with phenol-killed whole-cell S. typhimurium vaccine, respectively. The clinical status of the animals was followed and serial reisolation of vaccine and challenge strains from faeces was attempted. The immunization of calves with the live aro- auxotrophic S. typhimurium SL 1479 mutant proved to be unsuitable due to the death of calves after revaccination. The calves immunized with live auxotrophic gal E S. typhimurium CCM 3821 mutant proved to be protected against challenge with virulent S. typhimurium 4/5 strain administered orally at a dose of 10(6) colony forming units (CFU). The postvaccination complications showed serious shortcomings. The immunization of calves with three doses of whole-cell inactivated vaccine containing 5 strains of S. typhimurium was effective against oral challenge with virulent S. typhimurium 4/5 at a dose of 10(6) CFU.     

Use of a live attenuated Salmonella typhimurium vaccine to protect hens against Salmonella enteritidis infection while undergoing molt

Previous studies demonstrated that Salmonella enteritidis infections in hens undergoing molt via feed withdrawal were more severe than in full fed hens. We conducted two trials to determine if immunizing specific-pathogen-free, Salmonella-culture-negative hens via aerosol exposure to MeganVacl, a commercially available attenuated Salmonella typhimurium vaccine, would reduce transmission of S. enteritidis from infected hens to uninfected but contact-exposed hens during a molt. In trial 1, one group of hens received two aerosol doses of vaccine 2 wk apart whereas a second group of hens remained nonvaccinated. In trial 2, the vaccinated group received only one dose of vaccine. Two weeks after the final immunization, feed was removed from all the hens, and on day 4, the center hen in rows of 11 hens received a dose of 3 x 10(5) (trial 1) or 1.3 x 10(6) (trial 2). Transmission to the unchallenged hens was followed 3, 10, 17, and 24 days later. Vaccination reduced the horizontal spread of S. enteritidis in vaccinated hens compared with their nonvaccinated counterparts, with vaccinated hens shedding significantly less S. enteritidis on day 10 postchallenge in trial 1 and on days 3, 10, 17, and 24 in trial 2. Recovery of S. enteritidis from ovaries was significantly reduced in the vaccinated hens in trial 1 and from livers/spleens, ovaries, and cecum in trial 2. These studies indicate that immunization of hens with a live S. typhimurium vaccine could help reduce S. enteritidis problems during a molt situation.     

Development and duration of protection against salmonellosis in mice and sheep immunised with live aromatic-dependent Salmonella typhimurium.

The aim of this investigation was to determine the development and duration of protection in mice or sheep immunised with aromatic-dependent (aro-) Salmonella typhimurium strain CS332, by either parenteral or oral routes. Immunisation of mice by the intraperitoneal or sheep by the intramuscular routes was found to impart protection against oral challenge with the virulent parent S typhimurium strain CS94 as early as seven days after immunisation. In contrast, when immunisation was carried out by the oral route, protection was not evident until three weeks after immunisation. Regardless of the route of immunisation, mice were still partially protected at three months and were fully susceptible at six months after immunisation. In sheep, protection persisted for six months but not 12 months after immunisation. Only parenterally immunised mice and sheep developed high ELISA and, or, agglutinating antibody titres, and cutaneous delayed-type hypersensitivity (DTH) at three weeks after immunisation. Although both antibody and DTH were detectable three months after immunisation of mice with aro- S typhimurium strain CS332, none was detected at six months. Antibody measured by agglutination and ELISA was detectable six months after immunisation in sheep, although no DTH was evident. At 12 months after immunisation low levels of anti-LPS antibody (measurable by ELISA only) were detected in sheep immunised by the intramuscular route.     

Some safety aspects of salmonella vaccines for poultry: distribution and persistence of three Salmonella typhimurium live vaccines

The purpose of this study was to analyze the safety characteristics of three commercially available live Salmonella vaccine strains (vacT, Zoosaloral, and X3985) in relation to their persistence in individual animals but also within a flock and in the environment. In a first experiment, the digestive and systemic distributions in chickens were followed for 10 days in individually reared chickens that were orally inoculated at 1 day of age. Strain X3985 quickly disappeared from the digestive tract but remained in the liver until the end of this experiment, whereas strains vacT and Zoosaloral colonized the liver as well as the gut for 10 days. In the second trial, behavior of the vaccine strains was studied in groups of 20 chickens during 10 wk after a single oral administration to individual birds. Strain vacT remained in the environment of inoculated animals for 4-5 wk. Six weeks after the inoculation, vacT was not recovered from internal organs such as liver and spleen, and vacT disappeared from the digestive tract between the sixth and the 10th weeks. Comparatively, both Zoosaloral and X3985 vaccine strains persisted longer in the environment (8 wk at least). Of the vaccine strains, X3985 showed the greatest colonization of both systemic and digestive organs.     

Evaluation of protection against experimental salmonellosis in sheep immunised with 1 or 2 doses of live aromatic-dependent Salmonella typhimurium

The minimum number of doses of a live aromatic dependent (aro-) Salmonella typhimurium vaccine strain (SL1479), given by the intramuscular, oral or subcutaneous route required to protect sheep from experimentally-induced clinical salmonellosis, was determined. A significant reduction in mortalities and diarrhoea occurred in those sheep immunised with one or 2 intramuscular doses or 2 subcutaneous doses. On the other hand, sheep immunised with one subcutaneous dose were not protected. Immunisation with one or 2 oral doses also resulted in a significant reduction in mortality, although reduction in the prevalence of severe diarrhoea was less consistent. Sheep immunised with a single intramuscular dose of aro- S. typhimurium developed high levels of serum antibodies and significant delayed-type cutaneous hypersensitivity response to homologous Salmonella lipopolysaccharide and flagellin, whereas those with a single oral dose did not. It was concluded that immunisation of sheep with a single oral or intramuscular dose of live aro- S. typhimurium reduced mortalities and the prevalence of diarrhoea in sheep due to infection with virulent S. typhimurium.     

Growth of Salmonella typhimurium and Salmonella enteritidis in egg yolks from highly immunized hens.

This experiment was conducted to ascertain whether the growth of Salmonella Enteritidis (SE) or Salmonella Typhimurium (ST) would be suppressed in the presence of antibodies contained in egg yolks. Specific pathogen-free chickens (102 days of age) were subcutaneously immunized with oil-adjuvanted bacterin of SE or ST, twice within a four-week interval. During 160 to 170 days of age, eggs were collected, the yolks were removed and mixed with an equal volume of physiological buffered saline, inoculated with ten colony forming units (CFU) of SE or ST, and incubated at 37 degrees C or 20 degrees C for 23 hr. The growth of organisms in each yolk solution was examined. The egg yolk derived from non-immunized hens was examined in the same manner as the controls. There was no difference in the growth titer between the antibody-positive yolk and the negative yolk. The result suggests that the antibodies in the yolk do not influence the growth of each organism, even if the hens are highly immunized.     

Humoral responses and salmonellosis protection in chickens given a vitamin-dependent Salmonella typhimurium mutant

Humoral responses in chickens inoculated with an aromatic vitamin dependent (Aro-) Salmonella typhimurium mutant (STM) were studied to ascertain the efficacy of the organism as a vaccine for salmonellosis and possibly as a delivery system for antigens from enteric pathogens of chickens. Serum antibody responses in chickens that were given oral or subcutaneous inoculations of the bacterium followed the classic order of antibody production, with IgM being detected first, followed by IgG and IgA. Antibody responses in the gut of orally inoculated chickens were restricted to IgG and IgA. Weight gain measured in chickens given high doses of STM (up to 5 x 10(9)) orally, revealed that the bacterium did not adversely affect the chickens; in fact, inoculated chickens had significantly higher body weights than controls at the same age. Salmonellosis protection of chickens by oral vaccination with STM was examined in a vaccination/challenge experiment. The experiment revealed that oral vaccination reduced excretion of a virulent S. typhimurium used as the challenge organism.     

Detection of antibody to Salmonella enteritidis and S typhimurium in the yolk of hens' eggs

Enzyme-linked immunosorbent assays (ELISAs) have been developed to detect IgG antibodies to Salmonella enteritidis and S typhimurium in the yolk of hens' eggs. Better discrimination and more consistent results were obtained between eggs from experimentally infected and uninfected hens by using saline-dilution of yolk rather than chloroform extraction. Threshold absorbance values were determined in three salmonella-free flocks, and on the basis of these results ELISA optical density values greater than 0.25 were considered to be positive for antibodies to salmonella. Four flocks with a history of salmonella infection were examined; three contained birds which were seropositive for S enteritidis by ELISA and from which S enteritidis was isolated, and a large proportion of eggs from these birds contained antibody to S enteritidis. Eggs from the fourth flock had no detectable antibody, although serum antibody was detected in some birds. No salmonellae were isolated from the yolks of the eggs from any of the four flocks.     

Epizootiology of Salmonella typhimurium infection in chickens]

The incidence of S. typhimurium infections among fowl increased in thr region of Potsdam in general, and on various big farms in particular, 1976 and over the first half of 1977. The outbreaks included subclinical infections and clinically manifest diseases which caused remarkable loss of broilers from the affected stocks (up to 15.92 per cent). Parent stocks contaminated with S. typhimurium were to be the sources of infection in all cases. A total of 1,220 Salmonella strains were isolated from fowl and its environment, with 1,151 of them being S. typhimurium (2.98 per cent of all samples tested). The following amounts of S. typhimurium strains were isolated from different types of samples which had been collected from infected broiler stocks: 8.10 per cent from dead broilers, 5.86 per cent from dead broiler parents, 2.11 per cent from pulp linings of transport cages for day-old chicks, 1.23 per cent from litter, 1.0 per cent from hatching material (eggs or dead and jammed embryos, and 0.12 per cent from swabs used in hygiene supervision). No Salmonellae were isolated from feedstuff. The transmission of S. typhimurium, therefore, is though to have taken the route via the hatching egg and via congenitally infected chicks traded between breeders and propagation farms. The control and prophylaxis of S. typhimurium infections, therefore, should be based primarily on action in the centralised breeding stocks. Specific steps of such action are proposed. Fifty-three strains were biochemically and lysotypically analysed, with the following types being determined: ut/Ph 30 BT b, ut/Ph 30 BT c, n.c. 1/72/n.c. BT b, 2 n.c. BT a, and 1A/6 BT a. The first two types covered 84.9 per cent of all strains isolated from the fowl. All lysotype ut/Ph 30 strains isolated from fowl fell under the copenhagen variant which had rarely been isolated from man in the past. These results are likely to support the demand for a joint control programme for enteritis Salmonellae, with particular emphasis on S. typhimurium, for implementation in human and veterinary medicine.     

A new live Salmonella enteritidis vaccine for chickens--experimental evidence of its safety and efficacy

Within the works for the registration of a new live Salmonella Enteritidis vaccine for layers, safety and efficacy of the vaccine strain were tested by experimental studies. After oral administration of the single and the tenfold dose, no incompatibility reactions were seen in day-old chicks. The laying performance and the egg weight were not affected by the vaccination of the chickens during the laying period. There was only a limited period in which the excretion of the vaccine strain and its persistency in organs were seen. Even after the threefold oral vaccination the vaccine strain could not be isolated from eggs and internal organs of slaughtered chickens. Moreover, a high safety for non-target animals (cattle, pigs) could be established. Studies with BALB/c mice proved that a cell-mediated immunity and the development of complement-fixing antibodies is induced by the vaccine. Efficacy studies in target animals were carried out by a proved dependable oral challenge system that reproduces a latent infection with marked S. Enteritidis strains and by means of the seeder-bird method. The test results demonstrate that the vaccination is capable to avert or to reduce an infection significantly.     

Outbreak of Salmonella typhimurium in cats and humans associated with infection in wild birds

An outbreak of Salmonella typhimurium infection in cats and humans in Sweden in 1999, associated with wild birds, is described. In the county of Värmland, 62 sick cats were examined. All were anorectic and lethargic, 57 per cent had vomiting and 31 per cent had diarrhoea. It was considered likely that salmonellosis was transmitted from cats to humans, but there were only a few such cases.     

Colonization of a marker and field strain of Salmonella enteritidis and a marker strain of Salmonella typhimurium in vancomycin-pretreated and nonpretreated laying hens

This study was conducted to evaluate the influence of a vancomycin pretreatment on the ability of marker (nalidixic-acid resistant) Salmonella Enteritidis (SE(M)), field Salmonella Enteritidis (SE(E)), and marker Salmonella Typhimurium (ST(M)) strains to colonize within the intestinal and reproductive tracts and translocate to other organs of leghorn laying hens. In each of three trials, caged laying hens (76, 26, and 33 wk ofage) were divided into six groups designated to receive SE(M), SE(F), or ST(M), and half were pretreated with vancomycin (n = 11-12 hens). Vancomycin-treated hens received 10 mg vancomycin in saline/kilogram body weight orally for 5 days to inhibit Gram-positive bacteria within the intestines. On Day 6, all hens were concurrently challenged by oral, intravaginal, and intracolonal routes with Salmonella and placed into separate floor chambers by Salmonella strain. Two weeks postinoculation, all hens were euthanatized and the ceca, spleen, liver/gall bladder (LGB), upper (URT), and lower (LRT) reproductive tracts, and ovarian follicles were aseptically collected, and analyzed for Salmonella. Results did not differ for the three hen's ages and were therefore combined. The vancomycin pretreatment also had no significant effect on the colonization ability of SE(M), SE(F) or ST(M), and therefore results were combined within Salmonella strain. The marker strain of Salmonella Enteritidis was recovered from 21% of ceca, 4% of LGB, 9% of LRT, and 17% of the fecal samples. The field strain of Salmonella Enteritidis was recovered from 88% of ceca, 96% of spleen, 92% of LGB, 30% of LRT, 4% of URT, 13% of follicle, and 42% of the fecal samples. The marker strain of Salmonella Typhimurium was recovered from 100% of ceca, 74% of spleen, 91% of LGB, 30% of LRT, 9% of URT, 9% of follicle, and 100% of the fecal samples. Among ceca, spleen, LGB, and fecal samples, SE(F) and ST(M) colonization was significantly greater than SE(M) colonization. Overall prevalence of Salmonella in the reproductive tracts of challenged hens was relatively low, ranging from 4% to 30%.