Splenic leukocytes from chickens injected with Salmonella pullorum antigen stimulate production of corticosteroids by isolated adrenal cells

Corticosteroid levels in the blood serum of White Rock pullets were significantly increased within 1 hr after an iv injection of heat-killed Salmonella pullorum antigen (SP-Ag), and reached levels 7 to 10 times that of PBS-injected controls within 2 hr after SP-Ag injection. Incubations with isolated adrenal cells indicated that serum from SP-Ag-injected birds had the ability to stimulate the synthesis or release of corticosteroids twice that of serum PBS-injected birds. Stripping the serum from SP-Ag-injected birds with activated charcoal and precipitated silica (Quso G-32) removed the corticosteroids and the adrenal-stimulating ability. A 2-hr incubation of isolated adrenal cells with leukocytes from spleens removed from chickens 1 hr after injection with SP-Ag, using stripped serum as the medium, stimulated a two- to fivefold increase in corticosteroid as compared to splenic leukocytes from PBS-injected chickens incubated in the same medium. The results indicate that an "ACTH-like" substance was produced by the S. pullorum antigen-stimulated splenic leukocytes.     

Heterophils are associated with resistance to systemic Salmonella enteritidis infections in genetically distinct chicken lines

Heterophils mediate acute protection against Salmonella in young poultry. We evaluated susceptibility of genetically distinct lines of broilers to systemic Salmonella enteritidis (SE) infections. SE was administered into the abdomen of day-old chickens (parental lines [A and B]; F1 reciprocal crosses [C and D]) to assess modulation of leukocytes and survivability of chickens. Line A was more resistant to SE than line B; likewise cross D was more resistant than cross C. Significantly more heterophils migrated to the abdominal cavity post-infection in the resistant lines. These data indicate that increased heterophil influx to the infection site contributes to increased resistance against systemic SE infections in neonatal chickens.     

Epigenetic modification of TLRs in leukocytes is associated with increased susceptibility to Salmonella enteritidis in chickens

Toll-like receptors (TLRs) signaling pathways are the first lines in defense against Salmonella enteritidis (S. enteritidis) infection but the molecular mechanism underlying susceptibility to S. enteritidis infection in chicken remains unclear. SPF chickens injected with S. enteritidis were partitioned into two groups, one consisted of those from Salmonella-susceptible chickens (died within 5 d after injection, n = 6), the other consisted of six Salmonella-resistant chickens that survived for 15 d after injection. The present study shows that the bacterial load in susceptible chickens was significantly higher than that in resistant chickens and TLR4, TLR2-1 and TLR21 expression was strongly diminished in the leukocytes of susceptible chickens compared with those of resistant chickens. The induction of expression of pro-inflammatory cytokine genes, IL-6 and IFN-β, was greatly enhanced in the resistant but not in susceptible chickens. Contrasting with the reduced expression of TLR genes, those of the zinc finger protein 493 (ZNF493) gene and Toll-interacting protein (TOLLIP) gene were enhanced in the susceptible chickens. Finally, the expression of TLR4 in peripheral blood mononuclear cells (PBMCs) infected in vitro with S. enteritidis increased significantly as a result of treatment with 5-Aza-2-deoxycytidine (5-Aza-dc) while either 5-Aza-dc or trichostatin A was effective in up-regulating the expression of TLR21 and TLR2-1. DNA methylation, in the predicted promoter region of TLR4 and TLR21 genes, and an exonic CpG island of the TLR2-1 gene was significantly higher in the susceptible chickens than in resistant chickens. Taken together, the results demonstrate that ZNF493-related epigenetic modification in leukocytes probably accounts for increased susceptibility to S. enteritidis in chickens by diminishing the expression and response of TLR4, TLR21 and TLR2-1.     

QTL for resistance to Salmonella carrier state confirmed in both experimental and commercial chicken lines

The ability of chickens to carry Salmonella without displaying disease symptoms is responsible for Salmonella propagation in poultry stocks and for subsequent human contamination through the consumption of contaminated eggs or meat. The selection of animals more resistant to carrier state might be a way to decrease the propagation of Salmonella in poultry stocks and its transmission to humans. Five QTL controlling variation for resistance to carrier state in a chicken F₂ progeny derived from the White Leghorn inbred lines N and 6₁ had been previously identified using a selective genotyping approach. Here, a second analysis on the whole progeny was performed, which led to the confirmation of two QTL on chromosomes 2 and 16. To assess the utility of these genomic regions for selection in commercial lines, we tested them together with other QTL identified in an [N×6₁] × N backcross progeny and with the candidate genes SLC11A1 and TLR4 . We used a commercial line divergently selected for either low or high carrier-state resistance both in young chicks and in adult hens. In divergent chick lines, one QTL on chromosome 1 and one in the SLC11A1 region were significantly associated with carrier-state resistance variations; in divergent adult lines, one QTL located in the major histocompatibility complex on chromosome 16 and one in the SLC11A1 region were involved in these variations. Genetic studies conducted on experimental lines can therefore be of potential interest for marker-assisted selection in commercial lines.     

Disturbed immune-endocrine communication in autoimmune disease. Lack of corticosterone response to immune signals in obese strain chickens with spontaneous autoimmune thyroiditis

Antigenic challenge as well as injection of lymphokine-containing media lead to a transient increase of serum glucocorticoids, a phenomenon that has been implicated in the regulation of the specificity of immune responses. In the present study we examined the dialogue between the immune and the neuroendocrine systems in Obese strain (OS) chickens, an animal model for human Hashimoto thyroiditis. The following results were obtained: A) OS and normal White Leghorn (NWL) chickens, 5-mo-old, were immunized with sheep red blood cells followed by daily monitoring of corticosterone (CN) serum levels. Whereas in NWL animals CN serum levels markedly increase 3 to 4 days after immunization, OS animals did not respond with CN elevation. B) A single i.v. injection of conditioned medium (CM) from concanavalin A-stimulated spleen cells also led to a transient, dose-dependent peak in plasma CN (maximum after 30 min). This CN response to a given CM preparation was significantly lower in OS than in NWL animals. C) CM, whether obtained from OS or NWL splenocytes, were equally effective to stimulate CN production. D) A single i.v. injection of CM leads--concomitantly to the CN peak--to a decrease of the concanavalin A-mediated proliferative response of peripheral blood lymphocytes in both OS and NWL chickens. This suppression, however, was significantly more pronounced in NWL chickens. In summary, these data suggest a disturbance of the immune-neuroendocrine communication in OS chickens with spontaneous thyroid autoimmunity. The possible implications for the generation of "forbidden" autoimmune responses are discussed.     

In vivo and in vitro studies of genetic resistance to systemic salmonellosis in the chicken encoded by the SAL1 locus

A number of inbred lines of chickens have been shown to be resistant or susceptible to systemic salmonellosis caused by Salmonella enterica serovar Gallinarum in adult birds, or by S. enterica serovar Enteritidis and S. enterica serovar Typhimurium in young chicks. Resistant lines show only moderate pathology and low mortality rates, whereas susceptible lines display extensive pathological changes and higher levels of mortality following Salmonella infection. Genetic resistance to salmonellosis is dominant and not linked to sex, MHC or Slc11a1 (formerly known as Nramp1), which leads to resistance in mice and other species. A novel locus encoding resistance to salmonellosis has been identified on chicken chromosome 5, and designated SAL1. The nature of the differences in pathology found between resistant and susceptible chicken lines in vivo indicates that resistance is expressed at the level of the mononuclear phagocyte system. Macrophages from adult resistant line birds cleared Salmonella serovar Gallinarum from infected macrophages within 24 h, whereas Salmonella bacteria persisted within macrophages from susceptible line birds for at least 48 h. Clearance of Salmonella by macrophages was accompanied by a strong and reproducible respiratory burst response in resistant lines, but little or no response in susceptible lines. Macrophages from an outbred chicken line showed variable responses. No differences were seen in macrophage nitric oxide production in cells from resistant or susceptible lines. These differences suggest that increased macrophage antimicrobial activity correlates with resistance and that macrophage activity plays an important role in genetic resistance to systemic salmonellosis in the chicken.     

Bacteriophage therapy to reduce salmonella colonization of broiler chickens

Acute enteric infections caused by salmonellas remain a major public health burden worldwide. Poultry, particularly chickens, are known to be the main reservoir for this zoonotic pathogen. Although some progress has been made in reducing Salmonella colonization of broiler chickens by using biosecurity and antimicrobials, it still remains a considerable problem. The use of host-specific bacteriophages as a biocontrol is one possible intervention by which Salmonella colonization could be reduced. A total of 232 Salmonella bacteriophages were isolated from poultry farms, abattoirs, and wastewater in 2004 and 2005. Three phages exhibiting the broadest host ranges against Salmonella enterica serotypes Enteritidis, Hadar, and Typhimurium were characterized further by determining their morphology and lytic activity in vitro. These phages were then administered in antacid suspension to birds experimentally colonized with specific Salmonella host strains. The first phage reduced S. enterica serotype Enteritidis cecal colonization by > or = 4.2 log10 CFU within 24 h compared with controls. Administration of the second phage reduced S. enterica serotype Typhimurium by > or = 2.19 log10 CFU within 24 h. The third bacteriophage was ineffective at reducing S. enterica serotype Hadar colonization. Bacteriophage resistance occurred at a frequency commensurate with the titer of phage being administered, with larger phage titers resulting in a greater proportion of resistant salmonellas. The selection of appropriate bacteriophages and optimization of both the timing and method of phage delivery are key factors in the successful phage-mediated control of salmonellas in broiler chickens.     

Prevalence and antimicrobial resistance of Campylobacter spp. and Salmonella serovars in organic chickens from Maryland retail stores

Retail organic (n = 198) and conventional (n = 61) chickens were analyzed. Most organic (76%) and conventional (74%) chickens were contaminated with campylobacters. Salmonellae were recovered from 61% of organic and 44% of conventional chickens. All Salmonella enterica serovar Typhimurium isolates from conventional chickens were resistant to five or more antimicrobials, whereas most S. enterica serovar Typhimurium isolates (79%) from organic chickens were susceptible to 17 antimicrobials tested.     

Localization to chicken Chromosome 5 of a novel locus determining salmonellosis resistance

Clear genetic differences in the susceptibility of chickens to visceral infection by Salmonella have been observed and it has been possible to identify resistant and susceptible lines of inbred chickens. We report here the results of experiments to map directly the gene(s) controlling this trait in chickens by examining crosses between highly susceptible and highly resistant lines. In the mapping panel, a region on chicken Chromosome (Chr) 5 was found to have a large effect on resistance, and this effect was observed in three separate resource populations. Mapping of additional marker loci in the region of the resistance gene further localized it to a region of approximately 2 cM, close to the genes for creatine kinase (CKB) and dynein (DNCH1). This region shows conserved synteny with telomeric regions of human Chr 14 and mouse Chr 12. On the basis of this conserved synteny, this resistance gene seems unlikely to correspond to the previously identified salmonellosis resistance genes Lps (located on mouse Chr 4) or Nos(2) (located on mouse Chr 11). There was no association between Nramp1 and resistance in these crosses, although this gene was shown to contribute to resistance in other crosses. The homologous human and mouse regions at present contain no likely candidate genes for this trait. Thus this appears to be a novel resistance gene, which we designate SAL1.     

Immune Response and the B Blood Group Locus in Chickens

Chickens of three blood group genotypes, B1B1, B1B2 and B1B19, were compared in their ability to produce antibodies. The B locus is the major histocompatibility locus in this species. Homozygous B1 pullets had significantly higher adult mortality than did the B1 heterozygotes. In two separate experiments the B1B1 females produced significantly fewer antibodies to Salmonella pullorum than did the B1 heterozygotes. Also the B1B1 pullets responded with lower antibody titers following immunization with ferritin, bovine serum albumin and parainfluenza-3 virus, although the differences were not significant. The results of this study suggest that an immune response gene is associated with the major histocompatibility locus in chickens, paralleling the H-2 locus and Ir genes in mice.     

Dynamics of the avian inflammatory response to Salmonella following administration of the Toll-like receptor 5 agonist flagellin

Previous work has shown that flagellin (FGN) is a potent stimulator in vitro of phagocytic cell functions of chickens. The purpose of this study was to define the effects of FGN on the inflammatory response to Salmonella enteritidis (SE) in chickens. Intra-abdominal (IA) FGN administration caused significant increases in peripheral blood leukocytes (PBL) compared with SE-injected controls at 4 and 8 h postinjection (P相似文献   

SPF鸡盲肠复合微生态制剂代替抗生素预防肉仔鸡沙门茵感染

目的在雏鸡盲肠尽快建立完整健康菌群,通过“竞争排除（Competitiveexclusion,CE）”抑制病原菌定植感染。方法提取SPF鸡盲肠菌群,排除沙门菌感染的可能,制成复合微生态制剂,在鸡胚孵化19d啄壳时和21d雏鸡全出壳时进行两次喷雾接种,监测鸡群沙门菌抗体和粪便中沙门菌,测定成活率、生产性能。结果进行喷雾接种后的小鸡至12日龄可以完全不使用抗生素,成活率、采食量与体重略高于使用抗生素的对照组,能提高饲料报酬,饲养12、24、35d均未检出粪便沙门菌和血液沙门菌抗体,全程无沙门菌感染。结论SPF鸡盲肠复合微生态制剂能代替抗生素预防肉仔鸡沙门菌感染。     

Significance of the bacteriophage treatment schedule in reducing salmonella colonization of poultry

Salmonella remains the major cause of food-borne diseases worldwide, with chickens known to be the main reservoir for this zoonotic pathogen. Among the many approaches to reducing Salmonella colonization of broilers, bacteriophage offers several advantages. In this study, three bacteriophages (UAB_Phi20, UAB_Phi78, and UAB_Phi87) obtained from our collection that exhibited a broad host range against Salmonella enterica serovar Enteritidis and Salmonella enterica serovar Typhimurium were characterized with respect to morphology, genome size, and restriction patterns. A cocktail composed of the three bacteriophages was more effective in promoting the lysis of S. Enteritidis and S. Typhimurium cultures than any of the three bacteriophages alone. In addition, the cocktail was able to lyse the Salmonella enterica serovars Virchow, Hadar, and Infantis. The effectiveness of the bacteriophage cocktail in reducing the concentration of S. Typhimurium was tested in two animal models using different treatment schedules. In the mouse model, 50% survival was obtained when the cocktail was administered simultaneously with bacterial infection and again at 6, 24, and 30 h postinfection. Likewise, in the White Leghorn chicken specific-pathogen-free (SPF) model, the best results, defined as a reduction of Salmonella concentration in the chicken cecum, were obtained when the bacteriophage cocktail was administered 1 day before or just after bacterial infection and then again on different days postinfection. Our results show that frequent treatment of the chickens with bacteriophage, and especially prior to colonization of the intestinal tract by Salmonella, is required to achieve effective bacterial reduction over time.     

Novel function of the skin in calcium metabolism in female and male chickens (Gallus domesticus)

To study the role of the skin in differential calcium metabolism in White Leghorn chickens, we compared the composition of suction blister fluid (SBF) collected from cutaneous blisters with blood and serum in female and male animals in various physiological states. As an estimate for interstitial fluid (IF), SBF was used as a determinant of local cutaneous metabolism. Sample collection was carried out under ketamine–xylazine anesthesia. Eight chickens of both sexes were raised freely in similar environmental conditions and fed with similar food during their growth from juvenile to sexually mature and fully adult state. SBF, blood and serum were examined for concentrations of ionized Ca²⁺, Na⁺ and K⁺ with ion-selective electrodes (ISEs), and osmolalities by freezing point osmometry. pH and total protein content were also assessed. Our results showed that SBF of chickens is calcium-poor at the juvenile state and that it draws more Ca²⁺ in adult males than laying females of the same age. Interestingly, Ca²⁺ accumulation was observed also in females after laying had ceased. There was a positive correlation between blood and SBF Ca²⁺ in females but a negative one in males. In general, it was found that SBF of chickens was rich in Na⁺ and K⁺, was hypertonic compared to serum at the juvenile state and had a protein content of 36–47% of that in serum. Different from mammals, SBF in adult chickens was alkaline with the mean values of 8.7 ± 0.14 in females and 8.8 ± 0.06 in males. Age- and sex-related variability in cutaneous Ca²⁺ concentrations in chickens, and the differences of SBF composition between that of mammals point to a novel role of skin functions in avians. Possible functions of the skin as a dynamic calcium source balancing the free circulating Ca²⁺ levels and, also, as an excretory organ for Ca²⁺ are discussed.     

Production of germ line chimera by transfer of primordial germ cells in the domestic chicken (Gallus domesticus)

Blood was collected from Stage 13 to 14 (1) chick embryos. Primordial germ cells (PGCs) were separated from blood cells by Ficoll density gradient centrifugation. One hundred Rhode Island Red PGCs per embryo were transferred to the blood stream of Stage 14 to 15 White Leghorn embryos. Also, one hundred White Leghorn PGCs per embryo were transferred to the blood stream of Stage 14 to 15 Rhode Island Red embryos. Hatched male and female chicks were raised until sexual maturity, and progeny tests were performed by mating these PGC recipients with Rhode Island Red chickens of the opposite sex. Chicks apparently derived from the transferred PGCs, based on the feather color of the chicks, were produced from all 4 possible mating combinations. The present results indicate that the germ line of PGC recipient chickens consists of 2 distinct populations of germ cells.     

Spreading of<Emphasis Type="Italic">Salmonella enteritidis</Emphasis> in the cecum of chickens

Adhesion and colonization of high (2 x 10(8) CFU) and low doses (2 x 10(2) CFU) of Salmonella enteritidis (phage type 4) was determined in the ceca collected 6 h-4 weeks after inoculation (pi), of 1-d-old White Plymouth Rock orally-inoculated chickens. S. enteritidis was associated with the epithelial surface of the villi in the low-dose group 18 h-7 d pi, the penetration in the cecal lamina propria was observed on day 1 and 10 pi. In the high-dose group, adhesion and colonization was observed in all birds killed 6 h-14 d pi; penetration of the bacteria into the cecal lamina propria was seen 1-21 d pi. Large numbers of macrophage-like cells containing S. enteritidis were observed in the cecal lamina propria on days 3-21 pi. Colonization and migration by S. enteritidis in the intestinal tract of chickens was shown to be dose dependent.     

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 21 days of age. Groups of three birds were euthanised at intervals of 7 days (a total of 9 weeks) and one group was challenged with the same oocyst dose at 37 days p.i. and observed for 11 weeks. Blood samples were collected weekly, and sera were tested using IFAT. Chicken tissues were collected for PCR, quantitative PCR and immunohistochemistry. Two dogs approximately 45 days of age were fed with tissues from chickens euthanised at 138 and 159 days p.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 60 p.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 60 p.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 60 p.i. did not excrete N. caninum oocysts. This investigation indicates that chickens are resistant to experimental infection by N. caninum.     

Prevalence and Antimicrobial Resistance of Campylobacter spp. and Salmonella Serovars in Organic Chickens from Maryland Retail Stores

Retail organic (n = 198) and conventional (n = 61) chickens were analyzed. Most organic (76%) and conventional (74%) chickens were contaminated with campylobacters. Salmonellae were recovered from 61% of organic and 44% of conventional chickens. All Salmonella enterica serovar Typhimurium isolates from conventional chickens were resistant to five or more antimicrobials, whereas most S. enterica serovar Typhimurium isolates (79%) from organic chickens were susceptible to 17 antimicrobials tested.     

苏北地区白羽肉鸡父母代种鸡及其商品鸡中沙门氏菌的分离、鉴定与生物特性

【背景】沙门氏菌是一种重要的人兽共患食源性致病菌,控制沙门氏菌在动物中的感染和在食品生产链中的污染对于畜牧生产、公共卫生和食品安全具有重要意义。【目的】调查苏北地区某白羽肉鸡父母代种鸡及其商品鸡的沙门氏菌感染和流行情况,为其沙门氏菌防治提供参考依据。【方法】于2018?2019年从江苏泰州、宿迁、盐城、徐州、连云港等地区分别采集某白羽肉鸡父母代种鸡场产蛋鸡及其商品鸡场1?2日龄弱雏和10日龄雏鸡的泄殖腔拭子样品共360份,进行沙门氏菌的分离与鉴定,用纸片法和PCR方法对菌株耐药表型及相关耐药基因进行检测,通过多位点序列分型(multilocus sequence typing,MLST)技术对分离菌株的亲缘关系进行研究。【结果】从120份父母代产蛋鸡分离到3株沙门氏菌,从120份商品代1?2日龄弱雏分离到25株沙门氏菌,从120份商品代10日龄雏鸡分离到82株沙门氏菌。分离的沙门氏菌对多粘菌素B、亚胺培南、多西环素、甲氧苄啶、替加环素、氟苯尼考、头孢噻肟、恩诺沙星表现为全部敏感,对呋喃妥因、青霉素、利福平、利奈唑胺、万古霉素均表现为全部耐药;对阿莫西林耐药的菌株有30株(占27.27%),敏感的有80株(占72.73%)。耐药基因的PCR检测结果显示：β内酰胺类耐药基因blaTEM存在于所有沙门氏菌分离株中,但其他常见耐药基因在本研究的分离株中未发现。通过对分离菌7对管家基因进行PCR扩增、测序和MLST分型,发现所有沙门氏菌分离株的ST型均为ST11,属于肠炎沙门氏菌。MLST分型结果结合药敏试验结果表明,分离的沙门氏菌属于同一进化系统,具有相同或相似的来源。【结论】被调查商品鸡群的肠炎沙门氏菌感染很可能是其同一来源的父母代种鸡场垂直传播所致,这为该地区白羽肉鸡的沙门氏菌防治提供了参考。     

The Isolation of Salmonellae from the Bursa of Fabricius of Chickens Latently Infected with these Organisms

A nalidixic acid resistant mutant of Salmonella typhimurium was isolated more frequently from the Bursa of Fabricius than from the caecal contents, caecal tonsil, liver or spleen of chickens that had been given a culture of this mutant by mouth. Salmonellae were also isolated more frequently from the Bursa than from the caecal contents of naturally infected chickens.