Salmonella Heidelberg Research Articles

Effect of Infectious Bursal Disease (Ibd) Vaccines on Infection of Salmonella Heidelberg in Broiler Chickens

ABSTRACT Salmonella Heidelberg (SH) has represented a great concern to the Brazilian poultry industry in the last years. It is known that immunosuppression in poultry is a contributing factor to increase Salmonella faecal shedding and to disturb control programmes. Not only infectious bursal disease (IBD) virus but also some live vaccines have been reported to induce immunosuppression. In the present study we assessed the effects of two live vaccines against IBD on SH-infected broiler chicks. At 7 days of age, birds of three groups (vaccinated with recombinant HVT-IBD vector, with immune complex-IBD vaccine and unvaccinated) were orally challenged with 1 x 108 CFU of SH. A group of hatchmates remained unvaccinated/unchallenged to serve as negative controls. Caecal colonization and systemic invasion were evaluated by bacterial enumeration at 1, 3, 5, 7 and 14 days post-infection (Dpi) and SH faecal shedding assessed by cloacal swabs at 3, 7, 10 and 14 Dpi. The counts of SH in caecal contents were higher in birds vaccinated with immune complex-IBD than in those that received the HVT-IBD vector vaccine at 5, 7 and 14 dpi (p<0.01). There were no statistical differences in bacterial counts in liver and spleen among birds of different groups. Cloacal swabs also indicated that the birds vaccinated with immune complex-IBD shed more SH than those vaccinated with HVT-IBD vector or those unvaccinated (p<0.01). The results of the present study suggested that the immunosuppressive effect of the immune complex-IBD vaccine helped to increase the SH-faecal shedding in the infected birds.

Electrochemically-Activated Water Presents Bactericidal Effect Against Salmonella Heidelberg Isolated from Poultry Origin

Salmonella spp. are among the most important pathogens in poultry farming, and Salmonella Heidelberg (SH) is one of the most frequent serotypes isolated in Brazil. SH has a zoonotic potential and stands out as a pathogen that is difficult to eliminate from the poultry chain due to its resistance to disinfectants. One alternative to traditional disinfectants is the electrochemically-activated water (ECA), a bactericidal compound produced from the electrolysis of salt and water. ECA generators produce a compound that consists of free chlorine, hypochlorous acid, and other free radicals. This alternative control method is safe for human health and reduces environmental contamination. The present study aimed at evaluating the efficacy of ECA against 30 SH isolates from poultry origin in scenarios that simulated the chiller environment (4°C, 5 and 50 parts per million [ppm], 5 and 40 min of exposure) and the cleaning and disinfection process (25°C, 200 ppm, 5 and 10 min of exposure). In the quantitative test, SH was susceptible to ECA. The mean bacterial counts decreased significantly compared to the control group, especially at 200 ppm. At this concentration, ECA inhibited the growth of almost 87% of the Salmonella strains, and the results showed a significant decrease in the mean bacterial counts for both exposure times (5 and 10 min). These findings demonstrate that ECA is effective against SH in vitro and it is a possible alternative to disinfection in the poultry industry for the control of this pathogen. However, in situ tests in the food industry are needed.

Effect of Turkey-Derived Beneficial Bacteria Lactobacillus salivarius and Lactobacillus ingluviei on a Multidrug-Resistant Salmonella Heidelberg Strain in Turkey Poults

Effects of turkey-derived beneficial bacteria Lactobacillus ingluviei UMNPBX19 and Lactobacillus salivarius UMNPBX2 on Salmonella Heidelberg (SH) in turkey poults was investigated. Using in vitro studies, we determined each strain's resistance to pH 2.5 and 0.3% bile salts and their β-hemolysis activity. We also tested each strain's adherence to avian epithelial cells and exhibition of antimicrobial activity against major poultry-associated Salmonella. Moreover, using three in vivo experiments, we determined the effect of the strains in combination (LBIS) against SH in turkey poults. The treatment groups were negative control (-SH, -LBIS), SH control (+SH, -LBIS), and LBIS group (+SH, +LBIS). Supplementation of LBIS was done in drinking water throughout the study at a dose of 8 log CFU/gal. On day 7, poults were challenged with a 2011 ground turkey outbreak strain of SH at 5 × 105 CFU/mL, and the surviving pathogens were determined on day 7 postinoculation from the cecum, spleen, and liver. Both Lactobacillus strains exerted resistance to low pH and bile salts ( P < 0.05), showed adhesion to epithelial cells ( P < 0.05), but did not exhibit β-hemolysis. Cell-free culture supernatants of strains showed antimicrobial activity against Salmonella ( P < 0.05). Results from the in vivo studies revealed that LBIS significantly reduced dissemination of SH to the liver and spleen in all experiments, and colonization in the cecum in two of the three experiments (1.9- and 3.9-log CFU/g reductions), compared with the control. The results indicate that turkey-derived L. ingluviei UMNPBX19 and L. salivarius UMNPBX2 have potential beneficial effects against SH in turkeys. However, more studies to this effect are warranted.

Nanocomposite of Ag-Doped ZnO and AgO Nanocrystals as a Preventive Measure to Control Biofilm Formation in Eggshell and Salmonella spp. Entry Into Eggs.

Salmonella spp. is an important foodborne agent of salmonellosis, whose sources in humans often include products of avian origin. The control of this bacterium is difficult especially when Salmonella spp. is organized into biofilms. We hypothesized that the novel nanocomposites of ZnO nanocrystals doped with silver (Ag) and silver oxide (AgO) nanocrystals (ZnO:Ag-AgO) synthesized by the coprecipitation method could control or prevent the formation of Salmonella Enteritidis (SE) and Salmonella Heidelberg (SH) biofilm and its entry into turkey eggs. The diffraction characteristics of ZnO and AgO showed sizes of 28 and 30 nm, respectively. The Zn to Ag substitution into the ZnO crystalline structure was evidenced by the ionic radius of Ag+2 (1.26 Å), which is greater than Zn+2 (0.74 Å). For the SE analyses post-biofilm formation, the ZnO:Ag-AgO was not able to eliminate the biofilm, but the bacterial load was lower than that of the control group. Additionally, SE was able to infiltrate into the eggs and was found in both albumen and yolk. For the SH analyses applied onto the eggshells before biofilm formation, the ZnO:Ag-AgO treatment prevented biofilm formation, and although the bacterium infiltration into the eggs was observed in all treated groups, it was significantly smaller in ZnO:Ag-AgO pre-treated eggs, and SH could not reach the yolk. There was no difference in pore size between groups; therefore, the inhibition of biofilm formation and the prevention of bacterium entry into the egg were attributable to the use of ZnO:Ag-AgO, which was not influenced by the egg structure. Although the amount of Ag and Zn in the shell of the ZnO:Ag-AgO group was greater in relation to the control, this difference was not detected in the other egg components. In the search for new measures that are effective, safe and viable for controlling microorganisms in poultry farming, the application of a nanocomposite of Ag-doped ZnO and AgO nanocrystals appears as an alternative of great potential to prevent Salmonella sp biofilms in eggshells and other surfaces.

Transcriptome Analysis of Salmonella Heidelberg after Exposure to Cetylpyridinium Chloride, Acidified Calcium Hypochlorite, and Peroxyacetic Acid

The application of RNA sequencing in commercial poultry could facilitate a novel approach toward food safety with respect to identifying conditions in food production that mitigate transcription of genes associated with virulence and survivability. In this study, we evaluated the effects of disinfectant exposure on the transcriptomes of two field isolates of Salmonella Heidelberg (SH) isolated from a commercial broiler processing plant in 1992 and 2014. The isolates were each exposed separately to the following disinfectants commonly used in poultry processing: cetylpyridinium chloride (CPC), acidified calcium hypochlorite (aCH), and peroxyacetic acid (PAA). Exposure times were 8 s with CPC to simulate a poultry processing dipping station or 90 min with aCH and PAA to simulate the chiller tank in a poultry processing plant at 4°C. Based on comparison with a publicly available annotated SH reference genome with 5,088 genes, 90 genes were identified as associated with virulence, pathogenicity, and resistance (VPR). Of these 90 VPR genes, 9 (10.0%), 28 (31.1%), and 1 (1.1%) gene were upregulated in SH 2014 and 21 (23.3%), 26 (28.9%), and 2 (2.2%) genes were upregulated in SH 2014 challenged with CPC, aCH, and PAA, respectively. This information and previously reported MICs for the three disinfectants with both SH isolates allow researchers to make more accurate recommendations regarding control methods of SH and public health considerations related to SH in food production facilities where SH has been isolated. For example, the MICs revealed that aCH is ineffective for SH inhibition at regulatory levels allowed for poultry processing and that aCH was ineffective for inhibiting SH growth and caused an upregulation of VPR genes.

Targeting discriminatory SNPs in Salmonella enterica serovar Heidelberg genomes using RNase H2-dependent PCR

We report a novel RNase H2-dependent PCR (rhPCR) genotyping assay for a small number of discriminatory single-nucleotide polymorphisms (SNPs) that identify lineages and sub-lineages of the highly clonal pathogen Salmonella Heidelberg (SH). Standard PCR primers targeting numerous SNP locations were initially designed in silico, modified to be RNase H2-compatible, and then optimized by laboratory testing. Optimization often required repeated cycling through variations in primer design, assay conditions, reagent concentrations and selection of alternative SNP targets. The final rhPCR assay uses 28 independent rhPCR reactions to target 14 DNA bases that can distinguish 15 possible lineages and sub-lineages of SH. On evaluation, the assay correctly identified the 12 lineages and sub-lineages represented in a panel of 75 diverse SH strains. Non-specific amplicons were observed in 160 (15.2%) of the 1050 reactions, but due to their low intensity did not compromise assay performance. Furthermore, in silico analysis of 500 closed genomes from 103 Salmonella serovars and laboratory rhPCR testing of five prevalent Salmonella serovars including SH indicated the assay can identify Salmonella isolates as SH, since only SH isolates generated amplicons from all 14 target SNPs. The genotyping results can be fully correlated with whole genome sequencing (WGS) data in silico. This fast and economical assay, which can identify SH isolates and classify them into related or unrelated lineages and sub-lineages, has potential applications in outbreak identification, source attribution and microbial source tracking.

SALMONELLA TYPHIMURIUM VACCINE TO CONTROL A BRAZILIAN SALMONELLA HEIDELBERG STRAIN IN BROILER CHICKENS

Foodborne Salmonella infections in humans, which results from the consumption of contaminated poultry meat and eggs, are a major public health concern. Vaccination of animals against Salmonella is one strategy to prevent these infections and reduce the risks to public health. Live attenuated Salmonella enterica vaccines can confer protection against salmonellosis by inducing both cell-mediated and mucosal immune responses. This study assessed a live, attenuated Salmonella enterica Typhimurium (ST) vaccine in broiler chickens against a heterologous challenge with Salmonella Heidelberg (SH) by evaluating bacterial quantification, immune cells infiltration, and cytokine gene expression in the cecum. The treatments were: T1, non-vaccinated, non-challenged; T2, non-vaccinated, SH-challenged; T3, ST-vaccinated and SH-challenged. At 28 days of age, the ST-vaccinated group had significantly recovered reduction of SH in the crop (P<0,01) and cecum (P = 0,021) compared to the non-vaccinated SH-challenged group, with no significant changes (P˃0,05) in macrophages, T CD4+, or T CD8+ cells dynamics during the same period. Aerosol vaccination on the first day promoted greater interleukin-12 expression in the liver (P<0,.05) and interleukin-10 expression and T CD8+ cells in the ileum 16 hours after housing. After prime-boosted oral immunization on the 13th day, the vaccinated group had greater expression of macrophages and T CD4+ cells in the liver (P<0,05) than the control group. Two doses of a live ST-attenuated vaccine promoted a partial cross-protective effect against SH strain UFPR1 challenge in broilers.

Bacterial communication between Lactobacillus spp. isolated from poultry in the inhibition of Salmonella Heidelberg-proof of concept.

Bacterial communication has become an increasingly studied topic aiming at the discovery of new products to aid the treatment of diseases for which conventional options do not work. The production of safe foods, free of pathogens, has been receiving increasing attention due to market demands for food products of high quality and free of residues. This study assessed the communication between Lactobacillus spp. during the in vitro inhibition of Salmonella Heidelberg (SH) and the impact an autoinducer produced by a strain of Lactobacillus plantarum has on communication in the normal microbiota and inhibiting SH in newborn chicks. For this purpose, the isolates of Lactobacillus spp. were isolated cloacal swabs of broilers and identified through biochemical and molecular assays and were obtained from broiler farms. They later had their inhibitory potential against SH stimulated after contact with the autoinducer. For assessing bacterial communication (quorum sensing) during the inhibition of SH by Lactobacillus spp., a spot on the lawn assay was conducted. For the in vivo, 75 one-day-old chicks were divided in 5 experimental groups: control with no treatment; treatment with a Lactobacillus spp. pool on the first day; treatment with autoinducer on the second day; treatment with Lactobacillus spp. on the first day; and autoinducer on the second day and treatment without autoinducer. The autoinducer was assessed through an SH count in the ceca of the birds. The autoinducer produced by the strain of L. plantarum proved to be efficient for communicating with the other Lactobacillus spp. isolates as previous contact with SH induced the production of an autoinducer capable of increasing inhibition of SH both in vitro (in average 132.73%) and in vivo, acting similarly to the Lactobacillus spp. pool (probiotic) by decreasing the SH count in the ceca (64%-24h, 42%-96h, and 46%-168h).

Characterizing the Antimicrobial Function of a Dairy-Originated Probiotic, Propionibacterium freudenreichii, Against Multidrug-Resistant Salmonella enterica Serovar Heidelberg in Turkey Poults.

Antimicrobial potential of a dairy-origin probiotic bacteria, Propionibacterium freudenreichii, against multidrug-resistant Salmonella Heidelberg (SH) in turkey poults was determined in the current study. Employing in vitro experiments, two strains (subsp.) of P. freudenreichii: P. freudenreichii freudenreichii B3523 (PF) and P. freudenreichii shermanii B4327 (PS) were tested for their ability to resist low pH (2.5) and bile salts (0.3%). In addition, the ability of the strains to adhere to and invade avian epithelial cells was determined after exposure to Propionibacterium strains followed by SH challenge. Moreover, the antibacterial activity of the strains’ cell-free culture supernatants (CFCSs) were tested against three major foodborne pathogens, including SH. Furthermore, the susceptibility of the strains to common antibiotics used for human therapy was determined. The hemolytic properties of the strains were determined in comparison to Streptococcus pyogenes, a known hemolysis-causing pathogen. Appropriate controls were kept in all studies. Using two in vivo experiments, PF was tested against SH colonization of poult ceca and dissemination to liver and spleen. The four treatment groups were: negative control, PF control (PFC), SH control (SC), and a test group (PFS; PF + SH). The poults in the PFC and PFS groups were inoculated with 1010 CFU ml−1 PF on day 1 through crop gavage and subsequently supplemented through drinking water. On day 7, SC and PFS groups were challenged with SH at 106 CFU ml−1, and after 7 days, cecum, liver, and spleen were collected for determining surviving SH populations. Results indicated that both PF and PS resisted pH = 2.5 and 0.3% bile salts with surviving populations comparable to the control and adhered well onto the avian epithelial cell lines. The strains were susceptible to antibiotics and did not invade the epithelial cells or exhibit hemolytic properties. The CFCSs were highly bactericidal against all tested pathogens. In turkey poults, PF significantly reduced cecal colonization of SH and the dissemination of the pathogen to the liver, compared to the SH challenge controls (P < 0.05). Results revealed that PF, a non-host gastrointestinal tract-derived probiotic, could be an antibiotic alternative to prevent the early colonization of SH in poults, improving the preharvest safety of turkeys.

Lactobacillus bulgaricus, Lactobacillus rhamnosus and Lactobacillus paracasei Attenuate Salmonella Enteritidis, Salmonella Heidelberg and Salmonella Typhimurium Colonization and Virulence Gene Expression In Vitro.

Salmonella Enteritidis (SE), Salmonella Typhimurium (ST), and Salmonella Heidelberg (SH) have been responsible for numerous outbreaks associated with the consumption of poultry meat and eggs. Salmonella colonization in chicken is characterized by initial attachment to the cecal epithelial cells (CEC) followed by dissemination to the liver, spleen, and oviduct. Since cecal colonization is critical to Salmonella transmission along the food chain continuum, reducing this intestinal association could potentially decrease poultry meat and egg contamination. Hence, this study investigated the efficacy of Lactobacillus delbreuckii sub species bulgaricus (NRRL B548; LD), Lactobacillus paracasei (DUP-13076; LP), and Lactobacillus rhamnosus (NRRL B442; LR) in reducing SE, ST, and SH colonization in CEC and survival in chicken macrophages. Additionally, their effect on expression of Salmonella virulence genes essential for cecal colonization and survival in macrophages was evaluated. All three probiotics significantly reduced Salmonella adhesion and invasion in CEC and survival in chicken macrophages (p < 0.05). Further, the probiotic treatment led to a significant reduction in Salmonella virulence gene expression (p < 0.05). Results of the study indicate that LD, LP, and LR could potentially be used to control SE, ST, and SH colonization in chicken. However, these observations warrant further in vivo validation.

Salmonella Heidelberg: Genetic profile of its antimicrobial resistance related to extended spectrum β-lactamases (ESBLs).

The objective of this study was to evaluate the phenotypic and genotypic profile of antimicrobial susceptibility and the possible involvement of extended spectrum beta-lactamases (ESBLs) in the resistance profile of Salmonella Heidelberg (SH) isolated from chicken meat. We used 18 SH isolates from chicken meat produced in 2013 in the state of Paraná, Southern Brazil. The isolates were submitted to disk-diffusion tests and from these results it was possible to determine the number of isolates considered multiresistant and the index of multiple antimicrobial resistance (IRMA) against ten antimicrobials routinely used in human and veterinary medicine. It was considered multidrug resistant the isolate that showed resistance to three or more classes of antibiotics. Another test performed was the disc-approximation in order to investigate interposed zones of inhibition, indicative of ESBLs production. In the isolates that presented multidrug resistance (18/18), a search of resistance genes involved in the production of ESBLs was performed using PCR: blaCMY-2, blaSHV-1, blaTEM-1, blaCTX-M2, blaOXA-1, blaPSE-1 and AmpC. The overall antimicrobial resistance was 80.55%. The highest levels of resistance were observed for nalidixic acid and ceftiofur (100%). The most commonly resistance pattern found (42.1%) was A (penicillin-cephalosporin-quinolone-tetracycline). The results were negative for ghost zone formation, indicative of ESBLs. However, PCR technique was able to detect resistance genes via ESBLs where the blaTEM-1 gene showed the highest amplification (83.33%), and the second most prevalent genes were blaCMY-2 (38.88%) and AmpC gene (38.88%). The blaOXA-1 and blaPSE-1 genes were not detected. These results are certainly of concern since SH is becoming more prevalent in the South of Brazil and able to cause severe disease in immune compromised individuals, showing high antimicrobial resistance to those drugs routinely used in the treatment and control of human and animal salmonellosis.

Evaluation of a Probiotic and a Competitive Exclusion Product Inoculated In Ovo on Broiler Chickens Challenged with Salmonella Heidelberg

The present study evaluated a probiotic and a competitive exclusion product injected in ovo on day 18 of incubation together with Marek's disease vaccine in eggs of 56-week-old broiler breeders. Three experiments were carried out. The first trial evaluated the effect of treatments on hatchability, cecal colonization of Salmonella Heidelberg (SH), and intestinal mucosa immunity (immunoglobulin A levels in the intestinal fluid). The second trial evaluated the viability of the microorganisms in the products inoculated in a solution containing diluent and Marek's disease vaccine. The third trial evaluated the colonization of the cecal microbiota in non-challenged chickens during first four days of life by culturing cecal samples under aerobic and anaerobic conditions. Hatchability was not affected by the treatments. SH cecal counts were reduced in three-day-old broilers inoculated in ovo with the competitive exclusion product. Liver and spleen pool SH counts were not different among treatments. Broilers inoculated in ovo presented higher intestinal IgA titers 24 hours after SH challenge compared with the controls. When birds were not challenged, lower cecal microbial counts in aerobic culture were determined in the control group than in the probiotic group on day 3, and in the competitive exclusion group on day 2 when cultured in anaerobiosis. The products inoculated and diluted in the vaccine solution were viable at all analyzed periods when cultured in anaerobiosis. The results of this study suggest in-ovo inoculation is an effective route for the administration of the evaluated products, which effectively enhanced the broilers' immune response to a SH challenge, as shown by the increase in IgA titers, and the reduction in cecal Salmonella Heidelberg colonization with the in-ovo inoculation with the competitive exclusion product.

Risks Involved in the Use of Enrofloxacin for Salmonella Enteritidis or Salmonella Heidelberg in Commercial Poultry.

The objectives of the present study were to evaluate the risks involved in the use of Enrofloxacin for Salmonella Enteritidis (SE) or Salmonella Heidelberg (SH) in commercial poultry and determine the effects of a probiotic as an antibiotic alternative. Two experiments were conducted to evaluate the risks involved in the use of Enrofloxacin for SE or SH in commercial poultry. Experiment 1 consisted of two trials. In each trial, chickens were assigned to one of three groups; control + SE challenged; Enrofloxacin 25 mg/kg + SE; and Enrofloxacin 50 mg/kg + SE. Chickens received Enrofloxacin in the drinking water from days 1 to 5 of age. On day 6, all groups received fresh water without any treatment. All chickens were orally gavaged with 107 cfu/chick of SE at 7 days of age and euthanized on 8 days of age. In Experiment 2, turkey poults were assigned to one of the three groups; control + SH; probiotic + SH; and Enrofloxacin 50 mg/kg + SH. Poults received probiotic or Enrofloxacin in the drinking water from days 1 to 5 of age. On day 6, poults received fresh water without any treatment. Poults were orally gavaged with 107 cfu/poult of SH at 7 days of age. Poults were weighed and humanely killed 24 h post-SH challenge to evaluate serum concentration of fluorescein isothiocyanate-dextran to evaluate intestinal permeability, metagenomics, and SH infection. In both trials of Experiment 1, chickens treated with Enrofloxacin were more susceptible to SE organ invasion and intestinal colonization when compared with control non-treated chickens (P < 0.05). In Experiment 2, poults treated with 50 mg/kg of Enrofloxacin showed an increase in body weight, however, this group also showed an increase in SH susceptibility, intestinal permeability, and lower proportion of Firmicutes and Bacteroidetes, but with control group had the highest proportion of Proteobacteria. By contrast, poults that received the probiotic had the highest proportion of Firmicutes and Bacteroidetes, but lowest Proteobacteria. The results of the present study suggest that prophylactic utilization of Enrofloxacin at five times the recommended dose in poultry increases the susceptibility to salmonellae infections, and confirms that probiotics may be an effective tool in salmonellae infections.

Differential ex vivo responses of primary leukocytes from turkey pedigree lines to Salmonella Heidelberg

Escalating product recalls as a consequence of Salmonella-contaminated poultry products have resulted in detrimental economic impacts in the poultry industry. One potential long-term alternative method to Salmonella prevention is genetic selection to improve innate resistance. This study evaluated the ex vivo effects of Salmonella Heidelberg (SH) on phagocytic and bactericidal leukocyte function in turkeys from six pedigree lines (A-F). Day-of-hatch poults (n = 48) were placed and raised in cages (2 birds/gender/genetic line/cage) to 35 d when heterophils and peripheral blood mononuclear cells (PBMCs) were extracted from males and females of each line. Cells were used in phagocytic and bactericidal assays to determine the ex vivo effects of SH on turkey leukocyte activity. Data were analyzed using the Fit Model platform in JMP Pro 10.0 (SAS Institute Inc.) with differences considered significant at P ≤ 0.05 and data reported as LS Means with SEM. Although genetic line had no significant effect on phagocytosis of SH by heterophils and PBMCs, cumulatively, female cells exhibited higher phagocytosis potential than those from males. The main effect of gender was significant on bactericidal activity of PBMCs when incubated at a 1:10 and 1:100 PBMC to SH ratio. Genetic line also had a significant effect on bactericidal activity of PBMCs with cells from line F exhibiting the best activity. These results suggest that gender had a marked cumulative effect on phagocytosis of SH by heterophils and PBMCs while both genetic line and gender had a prominent effect on bacterial killing of SH by turkey PBMCs. Once able to determine genetic markers associated with these immune responses to Salmonella, genetic selection for increased resistance may become feasible in turkeys.

Effects of cold stress and Salmonella Heidelberg infection on bacterial load and immunity of chickens

We analysed the effects of cold stress (19 ± 1°C, 6 h /day, from the first to the seventh day of life) applied to specific pathogen free (SPF) chickens. On experimental Day 1 (ED1), chicks were divided into four groups: C (not infected and kept under thermoneutral condition); CS (not infected and cold stressed); PC (Salmonella Heidelberg (SH) infected and kept under thermoneutral condition) and PCS (SH infected and cold stressed). High concentrations of corticosterone were found in the cold stressed birds on ED7 and ED21, with a greater increase in birds of the PCS group. Stress or non-stressed SH-infected birds had high levels of norepinephrine on ED21. On ED21, an increased percentage and number of SH were found in birds of the PCS group. On ED7, a decrease in macrophages presenting MHCII, CD8+ and CD8+γδ cells was observed in the chickens of the CS group. Decrease was observed in CD3+ cells in the birds of the PCS group and increase in macrophages presenting MHCII cells and of the CD4+/CD8+ ratio in chickens of the CS group on ED21. There was a decrease in CD8+γδ cells in birds of the CS group on ED21 and in the CD3+ and CD8+cell numbers in chickens of the PCS group on ED21. Our results suggest that cold stress applied to chickens in the first 7 days of life increases both the hypothalamus pituitary adrenal axis and the sympathetic nervous system activities, leading to long-term immune cell dysfunction, thus allowing increased SH invasion and persistence within the birds’ body.

Effect of non-pharmaceutical compounds on shedding and colonization of Salmonella enterica serovar Heidelberg in broilers

The objectives of this study were to determine the effect of four non-pharmaceutical compounds on performance, mortality of broilers and their ability to reduce colonization and fecal shedding of Salmonella Heidelberg (SH) in broilers following SH challenge and feed withdrawal. Chicks were randomly assigned to water treatments containing organic acids (OA), essential oils (EO), lactic acid (LA), levulinic acid plus sodium dodecyl sulfate (L + SDS) or no added compounds (CON). Treatments were administered in drinking water on 0–7 and 35–42 days. One-half of the chicks were challenged with SH and placed in pens with unchallenged chicks on day one. Performance and mortality were determined during the 42-day study. Prevalence of SH was determined on drag swabs (0, 14 and 42 days) and in the ceca and crops (42 days). Broilers receiving EO had significantly (P < 0.05) greater weight gain and lower mortality than other treatments. Salmonella Heidelberg was absent from drag swabs on day 0, but present at 14 and 42 days. Challenged and unchallenged broilers receiving EO and LA had significantly (P < 0.05) lower SH in crops than other treatments. The essential oils and lactic acid used in the study may control SH contamination in crops of broilers when administered in drinking water.

Behavior of Salmonella heidelberg and Salmonella enteritidis strains following broiler chick inoculation: evaluation of cecal morphometry, liver and cecum bacterial counts and fecal excretion patterns

Over the years, Salmonella Heidelberg (SH) has gained prominence in North America poultry production and in the poultry production of other countries. Salmonella Heidelberg has been isolated and reported from poultry and poultry products in Brazil since 1962, whereas Salmonella Enteritidis (SE) has only emerged as a serious problem in poultry and public health since 1993. These strains of Salmonella can cause intestinal problems in newly hatched chicks, and infection may persist until adulthood. Upon slaughter of chickens, Salmonella can contaminate carcasses, a condition that poses a threat to human health. The aim of this study was to compare the fecal excretion of Salmonella Enteritidis and Salmonella Heidelberg in newly hatched chicks (orally inoculated with 105ufc/mL each) until 20 days of age. In addition, the ratio of cecal villus height:crypt depth (morphometry) and liver and cecum cell counts was analyzed in chicks ranging from 0 to 3 days of age and infected with these two Salmonella strains. One hundred seventeen chicks were separated into one of three experimental groups: a control group, an SE-infected group and an SH-infected group. Eight chicks per group were euthanized at 6, 12 and 72 hours post-inoculation (pi) to allow for Salmonella isolation from the liver and cecum and for the collection of the cecum for villi and crypt analysis. Other birds were allowed to mature to 20 days of age and cloacal swabs were taken at 2, 6, 13 and 20 days pi to compare the fecal excretion of inoculated strains. The Salmonella Enteritidis group had a higher number of cells excreted during the trial. Both strains were isolated from the liver and cecum by 6h pi. At 12h pi the Salmonella Heidelberg group had high cell counts in the cecum. No difference was found in liver cell counts. Both strains showed lower villus height:crypt depth ratio than the control group post-infection.

Efficacy of a Novel Trivalent Inactivated Vaccine Against the Shedding of Salmonella in a Chicken Challenge Model

The safety and efficacy of a novel trivalent inactivated Salmonella vaccine consisting of Salmonella Enteritidis (SE), Salmonella Typhimurium (ST), and Salmonella Infantis (SI) was evaluated with chickens. Chickens were injected subcutaneously with 0.5 ml per dose of the vaccine with an oil adjuvant. To assess the efficacy of the vaccine, vaccinated and unvaccinated chickens were challenged by oral inoculation of SE, ST, SI, and Salmonella Heidelberg (SH) virulent strains 4 wk after vaccination. The vaccinated chickens showed no clinical abnormalities, and the body weight gain after vaccination was comparable to that of unvaccinated chickens. The shedding of SE from the vaccinated chickens at 4, 7, 10, and 14 days postchallenge were significantly reduced in comparison with those of the unvaccinated chickens. The efficacy of the vaccine against ST and SI was comparable to that against SE. The shedding of SH at 4, 7, 10, and 14 days postchallenge was also significantly reduced in the vaccinated chickens compared with those in the unvaccinated suggesting that the vaccine could be effective against different serotype strains having O antigens homologous to those of the vaccine strains. These results suggest that the novel trivalent inactivated vaccine can be an effective tool for controlling the Salmonella infections of O4, O7, and O9 groups in chicken farms.