Immunogenic potential of a Salmonella Typhimurium live vaccine for pigs against monophasic Salmonella Typhimurium DT 193.

BACKGROUND: Monophasic Salmonella Typhimurium (mSTM) strains account for up to 8.6% of all human Salmonellosis cases. They have an increasing prevalence during recent years and several human cases with hospitalisation were reported. These strains are often isolated from pigs and pork - one primary source of human infection. A Salmonella Typhimurium (STM) live vaccine has been proven successful in controlling of STM infections in pigs for many years. The aim of this study was to test the immunogenicity of the vaccine in weaners during oral challenge with a virulent mSTM strain and to examine the kinetics of STM-specific IgA, IgM and IgG antibodies induced by vaccination and infection. RESULTS: Despite clinical signs being present in both groups, the vaccination led to a significant reduction of diarrhoea, overall clinical symptoms and a milder elevation of the body temperature. Necropsy revealed fewer pathological lesions in the gastrointestinal tract of vaccinated compared to control animals. Moreover, in the ileal and caecal mucosa and in the ileocaecal lymph nodes the challenge strain burden was significantly reduced by vaccination. Significant differences in the antibody responses of both groups were present during the vaccination period and after infection. In vaccinated animals Salmonella-specific IgA and IgG antibody levels increased significantly after vaccination and were even more pronounced in response to challenge. In contrast, similarly low levels of IgM antibodies were detected during the vaccination period in both vaccinated and non-vaccinated animals. However, after challenge IgM antibody levels increased significantly in control pigs while neither IgA nor IgG antibodies were detectable. CONCLUSION: The data demonstrate that mSTM can evoke clinical signs in weaners. Due to the vaccination their incidence and magnitude were significantly milder. Vaccination also led to a significantly reduced challenge strain burden in the intestine and the lymph nodes which is comparable to previous studies using the same vaccine in a challenge with biphasic STM. Therefore, it is concluded that this vaccine induces immunity against monophasic and biphasic STM strains. Furthermore, the results of antibody profiles in response to vaccination and infection provide additional evidence for humoral immune mechanisms triggered during Salmonella infection or vaccination.

Efficacy of Oral Rabies Vaccine Baits Containing SPBN GASGAS in Domestic Dogs According to International Standards.

(1) Background: The oral vaccination of free-roaming dogs against rabies has been developed as a promising complementary tool for mass dog vaccination. However, no oral rabies vaccine has provided efficacy data in dogs according to international standards. (2) Methods: To test the immunogenicity and efficacy of the third-generation oral rabies virus vaccine strain, SPBN GASGAS, in domestic dogs, dogs were offered an egg-flavoured bait containing 3.0 mL of the vaccine (107.5 FFU/mL) or a placebo egg-flavoured bait. Subsequently, these 25 vaccinated and 10 control animals were challenged approximately 6 months later with a dog rabies virus isolate. Blood samples were collected at different time points postvaccination and examined by ELISA and RFFIT. (3) Results: All but 1 of the 25 vaccinated dogs survived the challenge infection; meanwhile, all 10 control dogs succumbed to rabies. The serology results showed that all 25 vaccinated dogs seroconverted in ELISA (>40% PB); meanwhile, only 13 of the 25 vaccinated dogs tested seropositive ≥ 0.5 IU/mL) in RFFIT. (4) Conclusions: The SPBN GASGAS rabies virus vaccine meets the efficacy requirements for live oral rabies vaccines as laid down by the European Pharmacopoeia and the WOAH Terrestrial Manual. SPBN GASGAS already fulfilled the safety requirements for oral rabies vaccines targeted at dogs. Hence, the egg-flavoured bait containing SPBN GASGAS is the first oral vaccine bait that complies with WOAH recommendations for the intended use of oral vaccination of free-roaming dogs against rabies.

Pathogens Detected in 205 German Farms with Porcine Neonatal Diarrhea in 2017.

Neonatal diarrhea (ND) is still a frequently observed problem in modern industrial pig production. ND is predominantly caused by bacterial and viral pathogens. The objective of this study was to give an overview of different pathogens involved in ND in Germany. In 2017, a total number of 555 litters from 205 German pig farms with clinical ND were sampled with pooled fecal samples. All samples were analyzed regarding bacterial pathogens by culture and viral pathogens by polymerase chain reaction (PCR). Isolated strains of Clostridium (C.) perfringens, Escherichia (E.) coli, and C. difficile were further characterized by molecular techniques (e.g., PCR). There were 200 litters (36%), out of 555 sampled litters of 205 farms, which were positive for at least one, while most of them were positive for two or more pathogens. Toxin-producing C. perfringens type A could be detected in 122 farms (59.2%), C. difficile in 116 (56.1%), pathogenic E. coli in 79 (38.6%), and Rotavirus type A in 72 (35%). Among E. coli isolates, enterotoxigenic (8.8%) (F4 fimbriae positive (60.0%)) and necrotoxigenic E. coli (5.3%) were the most frequently detected pathotypes. In conclusion, in most of the farms with porcine ND it turned out to be a disease mainly caused by multiple pathogens, predominantly C. perfringens type A, pathogenic E. coli, and Rotavirus type A. Nevertheless, C. difficile and necrotoxigenic E. coli might be emerging pathogens in ND.

Invasion inhibition effects and immunogenicity after vaccination of SPF chicks with a Salmonella Enteritidis live vaccine.

OBJECTIVE: Meat and eggs from chickens infected with Salmonella Enteritidis, Salmonella Typhimurium and Salmonella Infantis are considered to be an important source of Salmonella infections for humans. In order to control Salmonella infections in chickens, basic biosecurity measures are taken in combination with inactivated or attenuated live vaccines. Apart from an adaptive immune response, some live vaccines also induce innate immune mechanisms that prevent or inhibit systemic invasion with homologous Salmonella serovars. It is unknown whether these invasion inhibition effects are also directed against heterologous Salmonella serovars. Furthermore, it is unclear whether the adaptive immune response after vaccination with a Salmonella Enteritidis phage type 4 live vaccine is also directed against other phage types of Salmonella Enteritidis and Typhimurium. MATERIAL AND METHODS: Specific pathogen-free day-old chicks were vaccinated orally with a commercially available Salmonella Enteritidis live vaccine. To test the invasion inhibition effect, the animals were challenged orally with a labelled Salmonella Typhimurium or Salmonella Infantis strain 1 day after vaccination. To demonstrate the adaptive immune response against non-phage type 4 Salmonella Enteritidis strains and a monophasic Salmonella Typhimurium strain, the chickens were challenged with Salmonella Enteritidis strains of phage types 1, 8 and 21 and a monophasic Salmonella Typhimurium strain (Definitive Type 193). After challenge, the abundance of the challenge strain in liver and cecal tissue was enumerated and compared with a corresponding control group. RESULTS: Findings showed that the live Salmonella Enteritidis vaccine inhibits systemic invasion after early infection with Salmonella Typhimurium and Salmonella Infantis. Furthermore, adaptive immunity against the tested non-phage type 4 Salmonella Enteritidis strains and the monophasic Salmonella Typhimurium strain was demonstrated. CONCLUSION AND CLINICAL RELEVANCE: The results of this study demonstrate that vaccination with the Salmonella Enteritidis phage type 4 live vaccine significantly inhibits the invasion of Salmonella Typhimurium and Infantis. Furthermore, an adaptive immune response was also detected against non-phage type 4 Salmonella Enteritidis strains and a monophasic Salmonella Typhimurium strain.

T-Cell Cytokine Response in Salmonella Typhimurium-Vaccinated versus Infected Pigs.

Vaccination with the live attenuated vaccine Salmoporc is an effective measure to control Salmonella Typhimurium (STM) in affected swine populations. However, the cellular immune response evoked by the Salmoporc vaccine including differences in vaccinated pigs versus non-vaccinated pigs upon STM infection have not been characterized yet. To investigate this, tissue-derived porcine lymphocytes from different treatment groups (vaccination-only, vaccination and infection, infection-only, untreated controls) were stimulated in vitro with heat-inactivated STM and abundances of IFN-γ, TNF-α and/or IL-17A-producing T-cell subsets were compared across organs and treatment groups. Overall, our results show the induction of a strong CD4+ T-cell response after STM infection, both locally and systemically. Low-level induction of STM-specific cytokine-producing CD4+ T cells, notably for the IFN-γ/TNF-α co-producing phenotype, was detected after vaccination-only. Numerous significant contrasts in cytokine-producing T-cell phenotypes were observed after infection in vaccinated and infected versus infected-only animals. These results suggest that vaccine-induced STM-specific cytokine-producing CD4+ T cells contribute to local immunity in the gut and may limit the spread of STM to lymph nodes and systemic organs. Hence, our study provides insights into the underlying immune mechanisms that account for the efficacy of the Salmoporc vaccine.

Vaccination and Infection of Swine With Salmonella Typhimurium Induces a Systemic and Local Multifunctional CD4+ T-Cell Response.

The gram-negative facultative intracellular bacteria Salmonella Typhimurium (STM) often leads to subclinical infections in pigs, but can also cause severe enterocolitis in this species. Due to its high zoonotic potential, the pathogen is likewise dangerous for humans. Vaccination with a live attenuated STM strain (Salmoporc) is regarded as an effective method to control STM infections in affected pig herds. However, information on the cellular immune response of swine against STM is still scarce. In this study, we investigated the T-cell immune response in pigs that were vaccinated twice with Salmoporc followed by a challenge infection with a virulent STM strain. Blood- and organ-derived lymphocytes (spleen, tonsils, jejunal and ileocolic lymph nodes, jejunum, ileum) were stimulated in vitro with heat-inactivated STM. Subsequently, CD4+ T cells present in these cell preparations were analyzed for the production of IFN-γ, TNF-α, and IL-17A by flow cytometry and Boolean gating. Highest frequencies of STM-specific cytokine-producing CD4+ T cells were found in lamina propria lymphocytes of jejunum and ileum. Significant differences of the relative abundance of cytokine-producing phenotypes between control group and vaccinated + infected animals were detected in most organs, but dominated in gut and lymph node-residing CD4+ T cells. IL-17A producing CD4+ T cells dominated in gut and gut-draining lymph nodes, whereas IFN-γ/TNF-α co-producing CD4+ T cells were present in all locations. Additionally, the majority of cytokine-producing CD4+ T cells had a CD8α+CD27- phenotype, indicative of a late effector or effector memory stage of differentiation. In summary, we show that Salmonella-specific multifunctional CD4+ T cells exist in vaccinated and infected pigs, dominate in the gut and most likely contribute to protective immunity against STM in the pig.

Application of an Endothelial Cell Culture Assay for the Detection of Neutralizing Anti-Clostridium Perfringens Beta-Toxin Antibodies in a Porcine Vaccination Trial.

BACKGROUND: Beta-toxin (CPB) is the major virulence factor of Clostridium perfringens type C, causing hemorrhagic enteritis in newborn pigs but also other animals and humans. Vaccines containing inactivated CPB are known to induce protective antibody titers in sow colostrum and neutralization of the CPB activity is thought to be essential for protective immunity in newborn piglets. However, no method is available to quantify the neutralizing effect of vaccine-induced antibody titers in pigs. (2) Methods: We developed a novel assay for the quantification of neutralizing anti-CPB antibodies. Sera and colostrum of sows immunized with a commercial C. perfringens type A and C vaccine was used to determine neutralizing effects on CPB induced cytotoxicity in endothelial cells. Antibody titers of sows and their piglets were determined and compared to results obtained by an ELISA. (3) Results: Vaccinated sows developed neutralizing antibodies against CPB in serum and colostrum. Multiparous sows developed higher serum and colostrum antibody titers after booster vaccinations than uniparous sows. The antibody titers of sows and those of their piglets correlated highly. Piglets from vaccinated sows were protected against intraperitoneal challenge with C. perfringens type C supernatant. (4) Conclusions: The test based on primary porcine endothelial cells quantifies neutralizing antibody activity in serum and colostrum of vaccinated sows and could be used to reduce and refine animal experimentation during vaccine development.

Demonstration of the efficacy of a Salmonella Enteritidis live vaccine for chickens according to the current European Pharmacopoeia Monograph.

Since 2013 the efficacy of new live Salmonella Enteritidis (SE) vaccines for chickens needs to be demonstrated according to European Pharmacopoeia Monograph 04/2013:2520 to receive approval in the EU. The purpose of this study was to determine whether a vaccine licensed since 1999 could also fulfil the required tests of the current guideline. For this, Salmonella-free chickens (n = 50) were vaccinated on their 2nd, 46th and 84th day of life with the live attenuated S. Enteritidis strain IDT No. 441/014. Non-vaccinated control animals (n = 50) were kept accordingly. To demonstrate the duration of immunity 20 animals of each group were challenge infected 65 weeks after the last vaccination with a virulent SE (PT 4) strain. According to the monograph, cloacal swabs were taken 3, 5, 7, 10 and 14 days post challenge (dpc). Tissue samples of liver, spleen, caeca, ovaries and oviduct were collected during necropsy of 10 animals per group on 7 and 14 dpc, respectively. All samples were analysed bacteriologically regarding the presence of the challenge strain. The number of challenge strain positive tissue samples and cloacal swabs was significantly reduced in vaccinated animals (p < 0.05). Therefore, the vaccine strain complied with the EP guideline. This study is the first that demonstrates the efficacy of this vaccine according to the current regulations. However, efficacy could also be shown during the development of the vaccine but by use of another animal model that comprised fewer animals per group. The use of this model is no longer accepted by EU regulatory authorities. The results need discussion in context with the 3R principle.

Vascular surgery of aortic thrombosis in a dog using Fogarty maneuver - technical feasibility.

Aortic thromboembolism is a rare and life-threatening disease in dogs. This report aims to describe the successful surgical treatment by use of a Fogarty Thrombectomy Catheter in an 8-year-old patient. The postsurgical intensive care therapy to prevent ischemia-reperfusion syndrome is specified, despite poor outcome in our case (owner elected euthanasia).

Expression of Toll-like receptors 2, 4 and 6 in equine endometrial epithelial cells: A comparative in situ and in vitro study.

Endometrial epithelial cells form a luminal barrier and are exposed to pathogens and non-infectious antigens. Toll-like receptors (TLRs) mediate pathogen defenses and tissue homeostasis, but are also involved in the pathogenesis of inflammatory and fibrosing alterations. Endometrial diseases are important causes of subfertility in mares. The pathogenesis of some types of persistent inflammation and periglandular fibrosis (endometrosis) is unknown. The aim of this study was to compare by in situ and in vitro immunohistochemistry the expression of TLRs 2, 4 and 6 in equine endometrial epithelial cells. An epithelial immunostaining for TLRs 2, 4 and 6 was detected in 76%, 57% and 90% of tissue sections, respectively. Positive cells lined the luminal surface, glandular ducts, mid glands and/or basal glands. An immunoreaction for TLRs 2, 4 and 6 was observed in 100%, 33% and 94% of cell cultures, respectively. The immunosignal was located in the cytoplasm and/or nucleus of endometrial epithelial cells under in situ and in vitro conditions. Results indicate a complex regulation of the epithelial expression of TLR 2, 4 and 6 proteins. The examined cell culture has to be regarded as suitable in vitro model. This study provides the basis for comparative investigations into the impact of different stimuli on the cellular expression of TLRs 2, 4 and 6. These will assist to find out if TLRs are involved in the pathogenesis of endometrial diseases and may help to understand as to why some mares develop persistent endometritis.

[Low-field magnetic resonance imaging of penetrating hoof injuries in 10 horses]. / Niederfeld-magnetresonanztomographische Untersuchung durchdringender Hufverletzungen bei 10 Pferden.

OBJECTIVE: The aim of the present study was to verify the diagnostic validity of low-field magnetic-resonance-imaging (MRI) findings in septic diseases of the foot region following penetrating sole injuries caused by nails. MATERIALS AND METHODS: MRI examinations were performed in 10 horses with injuries in the foot region. The diagnostic findings were validated by conventional methods (clinical, surgical, radiological, sonographic, and computed tomographic findings and/or post-mortem histological examination). RESULTS: Navicular bone involvement was revealed most accurately, with a high degree of predictability, and was best detected by fat-suppressed T2 or short-TI inversion recovery (STIR) sequences. MRI examinations of defects in the deep digital flexor tendon showed a high level of sensitivity, but these findings were less specific than changes to the navicular bone. They could be best reproduced in transverse T2-weighted fast spin echo sequences (T2w FSE). The penetration tract was recognisable in all cases and in all planes, and the T2w FSE proved to be very suitable for diagnosis. Septic bursitis was revealed least accurately by MRI. Bursal disease was best recognised in the sagittal plane. CONCLUSION AND CLINICAL RELEVANCE: MRI is a reliable method for confirming the diagnosis of diseases in the foot region after injuries caused by foreign bodies, particularly nails. A transverse T2w FSE is best suited for demonstrating a penetration tract and tendon damage. Visualisation of the penetration tract and secondary reactions of the navicular bone are crucial for diagnosing bursitis. Fat-suppressed sequences can clearly show bone involvement when the penetration tract has not reached the bone. The cases described illustrate that MRI is an appropriate method for evaluating puncture wounds in the foot region. Only MRI allows for intravital assessment of various structures within the hoof capsule. This information is essential for deciding upon targeted therapy while avoiding unnecessary therapies.