Salmonella enterica serovars in absence of ttrA and pduA genes enhance the cell immune response during chick infections.

Salmonella spp. is one of the major foodborne pathogens responsible for causing economic losses to the poultry industry and bringing consequences for public health as well. Both the pathogen survival ability in the intestinal environment during inflammation as well as their relationship with the host immune system, play a key role during infections in poultry. The objective of this study was to quantify the presence of the macrophages and CD4+/CD8+ cells populations using the immunohistochemistry technique, in commercial lineages of chickens experimentally infected by wild-type and mutant strains of Salmonella Enteritidis and Salmonella Typhimurium lacking ttrA and pduA genes. Salmonella Enteritidis ∆ttrA∆pduA triggered a higher percentage of the stained area than the wild-type, with exception of light laying hens. Salmonella Typhimurium wild-type strain and Salmonella Typhimurium ∆ttrA∆pduA infections lead to a similar pattern in which, at 1 and 14 dpi, the caecal tonsils and ileum of birds showed a more expressive stained area compared to 3 and 7 dpi. In all lineages studied, prominent infiltration of macrophages in comparison with CD4+ and CD8+ cells was observed. Overall, animals infected by the mutant strain displayed a positively stained area higher than the wild-type. Deletions in both ttrA and pduA genes resulted in a more intense infiltration of macrophages and CD4+ and CD8+ cells in the host birds, suggesting no pathogen attenuation, even in different strains of Salmonella.

The Efficiency of Bacteriophages Against Salmonella Typhimurium Infection in Native Noi Broilers

Though recently considered a therapeutic treatment for commercial broilers, little is known about the effects of bacteriophages on native, slow-growing birds. This study evaluated their efficacy against Salmonella enterica subsp. enterica serovar Typhimurium infected Noi chicken, a native Vietnamese broiler breed. In total, 420 birds were used in a completely randomized design consisting of seven treatments and four replicates of 15 birds. The treatments were NC (negative control), PC (positive control, S. Typhimurium challenged); NC+B1 and NC+B2 (negative control plus B1 or B2 bacteriophage, respectively); PC+B1, PC+B2 (positive control plus B1 or B2 bacteriophage, respectively) and PC+B1B2 (positive control plus both B1 and B2 bacteriophages). After four weeks of infection, the mortality rate in the PC group was 51.1% compared with 11.1% in the PC+B1B2 treatment. Bacteriophage administration had resulted in increased weight gain and decreased feed conversion ratio, particularly when both phages were included in the treatment (p<0.001). Moreover, the relative percentage of carcass weight was lowest in the PC treatment (66.9%) (p<0.001), whereas the other treatments registered similar carcass weight values. Regarding the internal organs, liver weight percentage was higher in the non-treated Salmonella group, and enlarged spleens were also noted in infected chickens even when treated with bacteriophages. The correlation between phage administration and blood parameters was unclear. Although the use of two bacteriophages for therapy was determined to be preferable for the majority of the criteria examined, further genetic characterization of the phages will be required before they can be widely used in chicken farms.(AU)

A histopathological and immunohistochemical study of experimental infected turkeys with a virulent Newcastle disease virus.

Newcastle disease (ND) is a highly contagious infection of many avian species, mainly chickens and turkeys, with a devastating impact on worldwide poultry production. This study was designed to examine the effect of virulent ND infection in turkey's tissues and the tissue tropism of the virus. During the previous study period, poults were inoculated at 32 days of age with 105 EID50 virulent Newcastle disease virus. Three poults on days 0, 1, 2, 3, 4, 6, 7, and 14 postinoculations (PI) were selected from each group. They were euthanized by intravenous sodium pentobarbital injection. After macroscopic observation, to histopathological and immunohistochemical studies, the spleen, bursa, cecal tonsils, intestine, proventriculus, lung, kidney, and brain were sampled. Clinically, the infected turkeys exhibited loss of appetite, severe depression, down on hock joint, white to greenish (sometimes bloody) diarrhea, nervous signs, and mild respiratory problems. Out of 45 birds inoculated, 9 (20%) died. Histopathological effects in lymphoid tissues included necrosis and penetration of mononuclear cells on day 4 PI, and subsequent follicular lymphoid depletion on days 6 and 8 PI was observed. Based on the immunohistochemical test, on day 3 in cecal tonsils and spleen, and on day 8 PI, all of them were positive for virus antigen. In conclusion, the NDV circulating in Iranian chicken flocks has the potential to cause severe illness in commercial turkeys.

Relevance of antibodies against the Chicken Anaemia Virus.

Chicken Infectious Anaemia (CIA) Virus (CAV) inhibits the function of multiple immune compartments. Mortality due to clinical infection is controlled in broilers by passive immunization derived from vaccinated breeders. Therefore, serological tests are often used in chicks to determine maternally-derived antibodies (MDA). We used a vaccine overdose-induced model of CIA. The model replicated the most common features of the disease. This model was used to determine the role of MDA in the protection of chicks. Hatchlings were tested for anti-CAV titers by ELISA and were sorted into groups based on antibody levels. SPF chicks were used as a no-antibody control. Lower specific antibody levels seemed to facilitate viral entry into the thymus, but viral levels, CD4+ and CD8+ counts, thymus architecture, and haematocrit were preserved by MDA, regardless of its levels. Levels of MDA are not correlated with protection from CIA, but are important for the progression CAV infection.

Immunological and bacteriological shifts associated with a flagellin-hyperproducing Salmonella Enteritidis mutant in chickens.

Salmonella Enteritidis causes infections in humans and animals which are often associated with extensive gut colonization and bacterial shedding in faeces. The natural presence of flagella in Salmonella enterica has been shown to be enough to induce pro-inflammatory responses in the gut, resulting in recruitment of polymorphonuclear cells, gut inflammation and, consequently, reducing the severity of systemic infection in chickens. On the other hand, the absence of flagellin in some Salmonella strains favours systemic infection as a result of the poor intestinal inflammatory responses elicited. The hypothesis that higher production of flagellin by certain Salmonella enterica strains could lead to an even more immunogenic and less pathogenic strain for chickens was here investigated. In the present study, a Salmonella Enteritidis mutant strain harbouring deletions in clpP and fliD genes (SE ΔclpPfliD), which lead to overexpression of flagellin, was generated, and its immunogenicity and pathogenicity were comparatively assessed to the wild type in chickens. Our results showed that SE ΔclpPfliD elicited more intense immune responses in the gut during early stages of infection than the wild type did, and that this correlated with earlier intestinal and systemic clearance of the bacterium.

A transient increase in MHC-IIlow monocytes after experimental infection with Avibacterium paragallinarum (serovar B-1) in SPF chickens.

Infectious coryza (IC), an upper respiratory tract disease affecting chickens, is caused by Avibacterium paragallinarum. The clinical manifestations of IC include nasal discharge, facial swelling, and lacrimation. This acute disease results in high morbidity and low mortality, while the course of the disease is prolonged and mortality rates are increased in cases with secondary infections. Studies regarding the immune response in infected chickens are scarce, and the local immune response is the focal point of investigation. However, a large body of work has demonstrated that severe infections can impact the systemic immune response. The objective of this study was to evaluate the systemic effects of Avibacterium paragallinarum (serovar B-1) infection on immune cells in specific pathogen-free (SPF) chickens. The current study revealed the presence of a transient circulating monocyte population endowed with high phagocytic ability and clear downregulation of major histocompatibility complex class II (MHC-II) surface expression. In human and mouse studies, this monocyte population (identified as tolerant monocytes) has been correlated with a dysfunctional immune response, increasing the risk of secondary infections and mortality. Consistent with this dysfunctional immune response, we demonstrate that B cells from infected chickens produced fewer antibodies than those from control chickens. Moreover, T cells isolated from the peripheral blood of infected chickens had a lower ability to proliferate in response to concanavalin A than those isolated from control chickens. These findings could be related to the severe clinical signs observed in complicated IC caused by the presence of secondary infections.

Modulation of the intestinal microbiota of broilers supplemented with monensin or functional oils in response to challenge by Eimeria spp.

The objective of this study was to evaluate the effect of supplementation with 100ppm sodium monensin or 0.15% of a blend of functional oils (cashew nut oil + castor oil) on the intestinal microbiota of broilers challenged with three different Eimeria spp. The challenge was accomplished by inoculating broiler chicks with sporulated oocysts of Eimeria tenella, Eimeria acervulina, and Eimeria maxima via oral gavage. A total of 864, day-old male broiler chicks (Cobb) were randomly assigned to six treatments (eight pens/treatment; 18 broilers/pen) in a 3 × 2 factorial arrangement, composed of three additives (control, monensin or blend), with or without Eimeria challenge. Intestinal contents was collected at 28 days of age for microbiota analysis by sequencing 16s rRNA in V3 and V4 regions using the Illumina MiSeq platform. Taxonomy was assigned through the SILVA database version 132, using the QIIME 2 software version 2019.1. No treatment effects (p > 0.05) were observed in the microbial richness at the family level estimated by Chao1 and the biodiversity assessed by Simpson's index, except for Shannon's index (p < 0.05). The intestinal microbiota was dominated by members of the order Clostridiales and Lactobacillales, followed by the families Ruminococcaceae, Bacteroidaceae, and Lactobacillaceae, regardless of treatment. When the controls were compared, in the challenged control group there was an increase in Erysipelotrichaceae, Lactobacillaceae, Bacteroidaceae, Streptococcaceae, and Peptostreptococcaceae, and a decrease in Ruminococcaceae. Similar results were found for a challenged group that received monensin, while the blend partially mitigated this variation. Therefore, the blend alleviated the impact of coccidiosis challenge on the microbiome of broilers compared to monensin.

Plant-based vaccine candidate against Infectious bursal disease: An alternative to inactivated vaccines for breeder hens.

Infectious bursal disease (IBD) is an acute, highly contagious immunosuppressive disease that affects young birds causing important economic losses in the poultry industry worldwide. Strict hygiene management together with effective vaccination programs are the most important strategies to prevent Infectious bursal disease virus entry in poultry production facilities. Hyperimmunisation of dams with inactivated vaccines just before the laying period provides passive immunity to the progeny that protects them during the critical first few weeks after hatching before vaccination with live attenuated virus takes place. In the present study, a safe and economic plant-based vaccine candidate against IBD intended for breeder hens was evaluated. We demonstrated that the recombinant immunogen is effective as booster for previously primed hens since it increases specific antibodies against VP2 that are transmitted to the offspring with titres and decay rate similar to those achieved by inactivated vaccine. Moreover, these maternally derived antibodies have virus neutralising activity and are able to confer protection against challenge in progeny, as evidenced by absence of bursal damage and low viral titres in this organ. Taking into account the disadvantages of inactivated vaccines as well as the benefits of plants as expression systems, such as time and cost efficiency, lower risk of contamination from animal pathogens and nearly unlimited scalability, a plant-based subunit IBD vaccine represents a viable alternative in the veterinary field.

Comparison between a commercial blend of functional oils and monensin on the performance and microbiota of coccidiosis-challenged broilers.

The aim of the study was to evaluate the effects of a cashew nut shell oil and commercial castor oil blend (CNSL-Castor oil) on the performance and microbiota of broiler chickens with and without coccidiosis challenge. A total of 864 one-day-old male chicks (Cobb) were randomly distributed to receive 6 treatments (8 pens/treatment; 18 chicks/pen) in a 3 × 2 factorial, with 3 additives (control [non-additives], 100 ppm sodium monensin, or 0.15% CNSL-Castor oil blend), and 2 levels of coccidiosis challenge at 14 D of age (unchallenged or inoculated by gavage with 1 mL of solution containing oocysts sporulated with Eimeria tenella, Eimeria acervulina, and Eimeria maxima). No differences in productive performance were observed among treatments in the pre-challenge period and in unchallenged birds (P > 0.05). Seven-days post-challenge, birds receiving monensin performed better than birds in the positive control group (non-additive and challenge) or in the CNSL-Castor oil group (P > 0.05). However, 14 D post-challenge, birds supplemented with CNSL-Castor oil presented higher weight gain and better feed conversion (P > 0.05), without any change in feed intake (P > 0.05). During the accumulated period (1 to 42 D of age), the live weight, weight gain, and feed intake did not differ between the CNSL-Castor oil and monensin groups, both of which presented higher values than the positive control. Lactobacillus spp. and Clostridium perfringens numbers were increased in the challenged birds (P < 0.05). CNSL-Castor oil supplementation reduced Clostridium cluster XIV, C. perfringens, and S. aureus, compared with the monensin and control groups (P > 0.05). In addition, the CNSL-Castor oil group presented the highest number of Lactobacillus spp. copies, followed by the monensin and positive control groups (P > 0.05). Thus, monensin and CNSL-Castor oil effectively minimized the impact of coccidiosis at different times. While monensin acts as an antimicrobial, CNSL-Castor oil modulates the intestinal microbiota with antimicrobial action against gram-positive bacteria, mainly C. perfringens and S. aureus.

Effect of functional oils on the immune response of broilers challenged with Eimeria spp.

Infection with Eimeria sp. results in the activation of multiple facets of the host immune system; the use of phytogenics can modulate the inflammatory response and improve the performance of the challenged animal. The aim of this study was to evaluate the effect of a commercial blend of cashew nut shell liquid (CNSL) and castor oil on the immune response of broilers challenged with coccidiosis. A total of 864 one-day-old male chicks (Cobb 500) were randomly distributed into six treatment groups (8 pens/treatment and 18 chicks/pen) in a three-by-two factorial design with three additives: control (non-additive), 100 ppm of monensin or 0.15% CNSL-castor oil. Challenge status was determined twice at 14 days of age. Unchallenged birds were inoculated by gavage with oocysts sporulated with Eimeria tenella, Eimeria acervulina and Eimeria maxima. Although the positive control (non-additive and challenged) and CNSL-castor oil treatment groups exhibited similar variation in weight gain (ΔBWG) compared to unchallenged birds fed without additives, the variation observed in birds fed diets containing CNSL-castor oil was associated with a higher maintenance requirement and not feed efficiency. In the second week after infection, ΔBWG of the CNSL-castor oil treatment group did not significantly change compared to the other treatment groups. At days 7 and 14 post-challenge, there was a higher excretion of oocysts in the control group, whereas the CNSL-castor oil and monensin groups did not differ. The CNSL-castor oil group exhibited increased gene expression of interferon (IFN), interleukin 6 (IL-6) and tumor necrosis factor (TNF), while the control group exhibited increased expression of cyclooxygenase (COX) and IL-1. The heterophils/lymphocyte ratio was low for the monensin treatment group. The unchallenged birds that received monensin treatment presented higher gene expression of IFN, COX and IL-1 compared to the other treatments, while the CNSL-castor oil group exhibited reduced gene expression, except for TNF. The commercial blend of cashew nut liquid and castor oil modulated the inflammatory response against Eimeria spp. In the absence of the parasite, there was no stimulation of genes involved in the inflammatory response, demonstrating that the blend is an effective tool in specifically modulating the immune system of birds afflicted with coccidiosis.

Memory immune responses and protection of chickens against a nephropathogenic infectious bronchitis virus strain by combining live heterologous and inactivated homologous vaccines.

In this study, we evaluated antibody and cell-mediated immune (CMI) responses in the mucosal and systemic compartments and protection against challenge with a nephropathogenic Brazilian (BR-I) strain of infectious bronchitis virus (IBV) in chickens submitted to a vaccination regime comprising a priming dose of heterologous live attenuated Massachusetts vaccine followed by a booster dose of an experimental homologous inactivated vaccine two weeks later. This immunization protocol elicited significant increases in serum and lachrymal levels of anti-IBV IgG antibodies and upregulated the expression of CMI response genes, such as those encoding CD8ß chain and Granzyme homolog A in tracheal and kidney tissues at 3, 7, and 11 days post-infection in the vaccinated chickens. Additionally, vaccinated and challenged chickens showed reduced viral loads and microscopic lesion counts in tracheal and kidney tissues, and their antibody and CMI responses were negatively correlated with viral loads in the trachea and kidney. In conclusion, the combination of live attenuated vaccine containing the Massachusetts strain with a booster dose of an inactivated vaccine, containing a BR-I IBV strain, confers effective protection against infection with nephropathogenic homologous IBV strain because of the induction of consistent memory immune responses mediated by IgG antibodies and TCD8 cells in the mucosal and systemic compartments of chickens submitted to this vaccination regime.

Evaluation of different live Salmonella enteritidis vaccine schedules administered during layer hen rearing to reduce excretion, organ colonization, and egg contamination.

Salmonellosis caused by Salmonella Enteritidis is a widespread zoonosis and poultry products are an important source of infection. This study was carried out to evaluate the protection of different vaccination schedules in layers using a live commercial attenuated Salmonella Enteritidis vaccine based on strain Sm24/Rif12/Ssq (AviPro® Salmonella Vac E, ELANCO) during rearing and egg production. Three hundred and fifty Salmonella-free chickens were distributed into 7 vaccinated groups and 1 unvaccinated group. Different vaccination schedules were performed combining either 1, 2, or 3 oral gavage doses. Chickens from Group A, B, and C were vaccinated once, either at the first day, at 7 or 16 wk old, respectively. Chickens from Group D were vaccinated twice-at the first day and 7 wk old. Chickens from Group E were vaccinated twice-at the first day and 16 wk old. Chickens from Group F were vaccinated twice-at 7 and 16 wk old. Chickens from Group G were vaccinated 3 times, following the manufacturer's recommendation: at the first day, 7 and 16 wk old. Chickens from Group H remained unvaccinated. Five challenge trials numbered 1 to 5 were carried out at 8, 12, 16, 29, and 55 wk old, respectively. After challenge, chickens were sampled by cloacal swabbing and, after euthanasia, livers, ovaries, spleens, and cecal contents were cultured to isolate S. Enteritidis. Additionally, eggs were collected after challenge and cultured to isolate S. Enteritidis on egg shells (Trials 4 and 5). Protection against experimental infection with a virulent nalidixic acid resistant S. Enteritidis strain K285/94, was evaluated by measuring reduction of excretion, colonization, invasion into organs, eggshell contamination, and egg production. The live S. Enteritidis vaccine protected the hens by reducing S. Enteritidis excretion, isolation from organs, and egg contamination. Higher protection throughout laying period was afforded after administration of three vaccine doses during rearing period.

Antigenicity, pathogenicity and immunosuppressive effect caused by a South American isolate of infectious bursal disease virus belonging to the "distinct" genetic lineage.

Infectious bursal disease virus (IBDV) is the causative agent of a highly contagious immunosuppressive disease affecting young chickens. The recently described "distinct IBDV" (dIBDV) genetic lineage encompasses a group of worldwide distributed strains that share conserved genetic characteristics in both genome segments making them unique within IBDV strains. Phenotypic characterization of these strains is scarce and limited to Asiatic and European strains collected more than 15 years ago. The present study aimed to assess the complete and comprehensive phenotypic characterization of a recently collected South American dIBDV strain (1/chicken/URY/1302/16). Genetic analyses of both partial genome segments confirmed that this strain belongs to the dIBDV genetic lineage and that it is not a reassortant. Antigenic analysis with monoclonal antibodies indicated that this strain has a particular antigenic profile, similar to that obtained in a dIBDV strain from Europe (80/GA), which differs from those previously found in the traditional classic, variant and very virulent strains. Chickens infected with the South American dIBDV strain showed subclinical infections but had a marked bursal atrophy. Further analysis using Newcastle disease virus-immunized chickens, previously infected with the South American and European dIBDV strains, demonstrated their severe immunosuppressive effect. These results indicate that dIBDV strains currently circulating in South America can severely impair the immune system of chickens, consequently affecting the local poultry industry. Our study provides new insights into the characteristics and variability of this global genetic lineage and is valuable to determine whether specific control measures are required for the dIBDV lineage. Research Highlights A South American strain of the dIBDV lineage was phenotypically characterized. The strain produced subclinical infections with a marked bursal atrophy. Infected chickens were severely immunosuppressed. The dIBDV strains are antigenically divergent from other IBDV lineages.

Arginine and manganese supplementation on the immune competence of broilers immune stimulated with vaccine against Salmonella Enteritidis.

This experiment was conducted to evaluate the combined effects of manganese-amino acid complex and arginine supplementation on the immune competence of broilers. On the day of hatch 640 male Cobb 500 broiler chicks assigned to two study groups (immune stimulate and non-stimulated). A 2 × 2 factorial arrangement of treatments was used with two manganese sources (MnSO4 or manganese-amino acid complex - MnAA) and two digestible Arg:Lys ratios (1.12 or 1.20). The treatments are: IM (80 ppm MnSO4); MnAA (40 ppm MnSO4 + 40 ppm MnAA); IM+Arg: 80 ppm MnSO4+ L-Arg (Arg:DigLys 1.20); MnAA+Arg: 40 ppm MnSO4 + 40 ppm MnAA + L-Arg (Arg:Lys 1.20). For treatments 1 and 2, the digestible Arg:Lys ratio was 1.12, considered normal for corn-soybean meal-based diets. Birds in the immune stimulated group received a dose of Salmonella Enteritidis vaccine. For growth performance and lymphoid organ development, no significant results were observed. Non-stimulated birds fed diets with Arg supplementation had higher percentage of mucosal T helper, T helper and T cytotoxic, compared to the normal Arg:Lys ratio (1.12). In the immune stimulated birds, broilers fed exclusive IM diet had a higher amount of T helper, T cytotoxic, activated T cytotoxic, and APC cells compared to broilers fed MnAA. The inorganic Mn diets, resulted in higher humoral antibody level (increased IgM levels) only when associated with supplementation of L-Arg. However, the use of an associated Mn source, could support high levels of IgM in commercial levels of Arg. No differences were observed to macrophage phagocytic activity analyses.

Cytokine gene transcription in the trachea, Harderian gland, and trigeminal ganglia of chickens inoculated with virulent infectious laryngotracheitis virus (ILTV) strain.

The objective of this study was to determine how cytokine transcription profiles correlate with patterns of infectious laryngotracheitis virus (ILTV) replication in the trachea, Harderian gland, and trigeminal ganglia during the early and late stages of infection after intratracheal inoculation. Viral genomes and transcripts were detected in the trachea and Harderian gland but not in trigeminal ganglia. The onset of viral replication in the trachea was detected at day one post-infection and peaked by day three post-infection. The peak of pro-inflammatory (CXCLi2, IL-1ß, IFN-γ) and anti-inflammatory (IL-13, IL-10) cytokine gene transcription, 5 days post-infection, coincided with the increased recruitment of inflammatory cells, extensive tissue damage, and limiting of virus replication in the trachea. In contrast, transcription of the IFN-ß gene in the trachea remained unaffected suggesting that ILTV infection blocks type I interferon responses. In the Harderian gland, the most evident transcription change was the early and transient upregulation of the IFN-γ gene at 1 day post-infection, which suggests that the Harderian gland is prepared to rapidly respond to ILTV infection. Overall, results from this study suggest that regulation of Th1 effector cells and macrophage activity by Th1/2 cytokines was pertinent to maintain a balanced immune response capable of providing an adequate Th1-mediated protective immunity, while sustaining some immune homeostasis in preparation for the regeneration of the tracheal mucosa.

Experimental Neospora caninum infection in chickens (Gallus gallus domesticus) with oocysts and tachyzoites of two recent isolates reveals resistance to infection.

The importance of birds in the biological cycle of Neospora caninum is not clear. We report unsuccessful Neospora infection in chickens (Gallus gallus domesticus) using two isolates of N. caninum. In experiment #1, 30 White Leghorn chickens were orally inoculated with viable N. caninum oocysts (NC-SP1 isolate, 200 oocysts per bird) via the crop at 21days of age. Groups of three birds were euthanised at intervals of 7days (a total of 9weeks) and one group was challenged with the same oocyst dose at 37daysp.i. and observed for 11weeks. Blood samples were collected weekly, and sera were tested using IFAT. Chicken tissues were collected for PCR, quantitative PCR and immunohistochemistry. Two dogs approximately 45days of age were fed with tissues from chickens euthanised at 138 and 159daysp.i. The results indicated that the chickens were resistant to neosporosis as revealed by failure to seroconvert, to detect parasite DNA or N. caninum antigen by immunohistochemistry in inoculated bird tissues, and by no oocyst excretion by the dogs fed avian tissues. Similar results were obtained in experiment #2, in which 34 1-week-old chickens were each s.c. inoculated with 100,000 tachyzoites of the NcWTDMn1 isolate of N. caninum. The chickens were euthanised on days 7, 15, 22, 28, 36 and 60p.i. At necropsy, all tissues and serum from each bird were collected. All chickens remained asymptomatic, and N. caninum antigen was not detected by immunohistochemistry. Seven chickens euthanised at day 60p.i. demonstrated low (1:25 dilution) levels of antibodies by using the Neospora agglutination test. Two 12-week-old dogs fed tissues pooled from 10 inoculated chickens euthanised at day 60p.i. did not excrete N. caninum oocysts. This investigation indicates that chickens are resistant to experimental infection by N. caninum.

Evaluation of recombinant Salmonella vaccines to provide cross-serovar and cross-serogroup protection.

Historically, Salmonella vaccines have been either live attenuated or killed bacterin vaccines that fail to offer cross-serogroup protection, which limits risk mitigation and protection of consumers. Subunit recombinant vaccines which possess highly conserved antigens offer potential to provide cross-serogroup protection, and the ability to express immune-enhancing molecules that promote recognition by the immune system. Six Salmonella subunit vaccine candidates were developed in either attenuated S. Enteritidis (SE) or S. Typhimurium (ST) that cell-surface express antigenic epitopes of high mobility group box 1 immune-enhancing sequence (H), peptidoglycan associated lipoprotein (P), and Omp18 protein Cj0113 (C) in different pattern arrangements for evaluation against S. Heidelberg (SH) challenge in broilers. In exp. 1, chicks were orally vaccinated with SE-CPH, SE-HCP, SE-CHP, ST-CPH, ST-HCP, or ST-CHP at 1 × 107 cfu/chick, or saline on d 1 and d 14. On d 17 all birds were challenged with 6 × 106 cfu/chick SH, and ceca collected on d 23 and d 28. On d 23 only SE-CPH reduced (P < 0.05) SH recovery at 0.34 ± 0.23 log10 cfu when compared to control at 1.19 ± 0.26 log10 cfu. On d 28, SE-CPH and ST-HCP reduced SH recovery at 0.40 ± 0.40 and 0.51 ± 0.26 log10 cfu, respectively in comparison to control at 1.36 ± 0.23 log10 cfu. For exp. 2, chicks were orally vaccinated with 1 × 108 cfu/chick SE-CPH, SE-HCP, SE-CHP or saline on d 1. At d 7 all chicks were orally challenged with 7 × 106 cfu/chick SH and ceca collected on d 28 and d 35. SE-CPH reduced (P < 0.05) SH recovery on d 28 when compared to control (6.16 ± 0.13 vs. 4.71 ± 0.55 log10 cfu). In exp 3, chicks were vaccinated by spray in a commercial vaccination cabinet with SE-CPH vaccination, 1.6 × 107 cfu/chick, or saline. Birds were challenged on d 14 with 3 × 107 cfu/chick SH and ceca collected on d 18 and d 25. SE-CPH reduced SH recovery (P < 0.05) on d 18, 2.75 ± 0.05 log10 cfu, and d 25, 1.89 ± 0.43 log10 cfu, as compared to control chickens at 5.6 ± 0.37 (d 18) and 3.98 ± 0.5 log10 cfu (d 25). The results of these experiments suggest that cross-serogroup protection is possible using these SE and ST-vectored subunit vaccines.

A recombinant avian antibody against VP2 of infectious bursal disease virus protects chicken from viral infection.

A stable cell-line was established that expressed the recombinant avian antibody (rAb) against the infectious bursal disease virus (IBDV). rAb exhibited neutralization activity to IBDV-B87 strain in DF1 cells. The minimum rAb concentration required for inhibition of the cytopathic effect (CPE) was 1.563µg/mL. To test the efficacy of rAb, a 168-h cohabitation challenge experiment was performed to transmit the disease from the chickens challenged with vvIBDV (HLJ0504 strain) to three test groups of chickens, i.e. (1) chickens treated with rAb, (2) chickens treated with yolk antibody, and (3) non-treatment chickens. The survival rates of chickens treated with rAb, yolk antibody and without treatment were 73%, 67% and 20%, respectively. Another batch of chickens was challenged with IBDV (BC6/85 strain) and then injected with rAb (1.0mg/kg) 6, 24 and 36h post-challenge. Non-treatment chickens had 100% morbidity, whereas those administered with rAb exhibited only 20% morbidity. Morbidity was evaluated using clinical indicators and bursal histopathological section. This study provides a new approach to treating IBDV and the rAb represents a promising candidate for this IBDV therapy.

Early immune responses and development of pathogenesis of avian infectious bronchitis viruses with different virulence profiles.

Avian infectious bronchitis virus (IBV) primarily replicates in epithelial cells of the upper respiratory tract of chickens, inducing both morphological and immune modulatory changes. However, the association between the local immune responses induced by IBV and the mechanisms of pathogenesis has not yet been completely elucidated. This study compared the expression profile of genes related to immune responses in tracheal samples after challenge with two Brazilian field isolates (A and B) of IBV from the same genotype, associating these responses with viral replication and with pathological changes in trachea and kidney. We detected a suppressive effect on the early activation of TLR7 pathway, followed by lower expression levels of inflammatory related genes induced by challenge with the IBV B isolate when compared to the challenge with to the IBV A isolate. Cell-mediated immune (CMI) related genes presented also lower levels of expression in tracheal samples from birds challenged with B isolate at 1dpi. Increased viral load and a higher percentage of birds with relevant lesions were observed in both tracheal and renal samples from chickens exposed to challenge with IBV B isolate. This differential pattern of early immune responses developed after challenge with IBV B isolate, related to the downregulation of TLR7, leading to insufficient pro-inflammatory response and lower CMI responses, seem to have an association with a most severe renal lesion and an enhanced capability of replication of this isolate in chicken.

Experimental infection of chickens by a flagellated motile strain of Salmonella enterica serovar Gallinarum biovar Gallinarum.

Salmonella enterica subsp. enterica serovar Gallinarum biovar Gallinarum (SG) causes fowl typhoid (FT), a septicaemic disease which can result in high mortality in poultry flocks. The absence of flagella in SG is thought to favour systemic invasion, since bacterial recognition via Toll-like receptor (TLR)-5 does not take place during the early stages of FT. In the present study, chicks susceptible to FT were inoculated with a wild type SG (SG) or its flagellated motile derivative (SG Fla(+)). In experiment 1, mortality and clinical signs were assessed, whereas in experiment 2, gross pathology, histopathology, systemic invasion and immune responses were evaluated. SG Fla(+) infection resulted in later development of clinical signs, lower mortality, lower bacterial numbers in the liver and spleen, and less severe pathological changes compared to SG. The CD8(+) T lymphocyte population was higher in the livers of chicks infected with SG at 4 days post-inoculation (dpi). Chicks infected with SG had increased expression of interleukin (IL)-6 mRNA in the caecal tonsil at 1 dpi and increased expression of IL-18 mRNA in the spleen at 4 dpi. In contrast, the CD4(+) T lymphocyte population was higher at 6 dpi in the livers of birds infected with SG Fla(+). Therefore, flagella appeared to modulate the chicken immune response towards a CD4(+) T profile, resulting in more efficient bacterial clearance from systemic sites and milder infection.