Pathological changes in the immune organs of broiler chickens fed on corn naturally contaminated with aflatoxins B1 and B2

The purpose of this study was to evaluate the underlying basis for aflatoxin-induced immunosuppression in the broiler chicken by detecting pathological lesions and apoptosis in thymus, bursa of Fabricius (BF) and spleen. COBB500? male broiler chicks were randomly allocated to two groups. The control group was fed on a basal corn-based diet while the other group (the AFB group) was fed on a similar diet but the corn was naturally contaminated with aflatoxins B₁ and B₂. Histopathological examination revealed that in the AFB group there was more nuclear debris in the three immune organs and obvious congestion of red pulp in the spleen, when compared with the control group. Ultrastructural examination showed lesions in the lymphocytes and reticulocytes of the three immune organs, the mucosal epithelium of the BF and the plasmocytes of the spleen. Increased apoptotic cells and an impaired membrane system (including nuclear membrane, mitochondria and endoplasmic reticulum [ER]) could be observed in the three immune organs in birds of the AFB group. In the plasmocytes, dilated rough endoplasmic reticulum contained electron-dense matrix. By flow cytometry, the percentages of apoptosis were significantly higher (P < 0.01) in the three organs of the AFB group than those of the control group. These observations suggested that the lesions of the immune organs were related to the immunosuppression, and that the apoptosis might be initiated by the mitochondrial pathway and ER chaperone pathway.     

Antimicrobial resistance and epidemiology of Escherichia coli in broiler breeder chickens

Escherichia coli isolates were obtained from two flocks of broiler breeder chickens beginning at 1 day old. Antimicrobial sensitivities were determined and isolates were grouped on the basis of their antibiogram patterns. In both flocks there was an initial dramatic shift in the antimicrobial resistance patterns of the E. coli isolates which changed from sensitive to multiresistant. Both flocks were given spectinomycin in drinking water during the first 3 days on the rearer farm. Statistical analysis of the E. coli isolates in Flock 2 revealed that there was a significant difference between E. coli obtained from 1-day-old birds and those obtained from 1-week-old birds in terms of the proportion of isolates that were resistant to spectinomycin. It is possible that the use of spectinomycin selected for resistant E. coli isolates which became dominant in the flocks soon after treatment. There was a strong association between resistance to spectinomycin and resistance to other antimicrobial agents, in particular, suiphafurazole and chloramphenicol.     

Avian pneumovirus infection in broiler chicks inoculated with Escherichia coli at different time intervals

The effects of dual infection of 1-day-old broiler chicks with a chicken isolate of avian pneumovirus (APV) and a pool of pathogenic Escherichia coli strains were studied by supraconjunctival application of the bacteria simultaneously with the virus, or at 4, 7 or 11 days afterwards. When the agents were given together, the clinical disease was significantly more severe than that caused by the virus alone, but when the bacterium was given later the signs were less severe. None of the infections resulted in swollen head syndrome by 32 days. All mixed infections caused moderate to severe congestion in the turbinates, when birds were examined at 32 days of age, at which time no such lesions were present in birds having been infected with APV alone. E. coli was isolated from almost 100% of birds with mixed infections, while rates of those given only E. coli isolation varied between 56 and 67%. Furthermore, E. coli colony counts were consistently higher from mixed infection groups. Virus persistence in the choanal cleft was slightly prolonged in birds with the simultaneous mixed infection. Although the pool of E. coli included O2, O78 and O18 serotypes, only those of the O2 serotype and a small number of untypable strains were re-isolated from selected mixed and single E. coli-infected groups. Mixed APV and E. coli infection did not affect APV enzyme-linked immunosorbent assay antibody titres at 21 or 32 days. Thus, experimental infection of broiler chicks with APV and E. coli, simultaneously or at intervals afterwards, demonstrated a synergistic effect between the two agents, but none of the infection protocols caused swollen head syndrome.     

Individual and combined effects of ochratoxin A and Salmonella enterica serovar Gallinarum infection on pathological changes in broiler chickens

A study was conducted to evaluate the individual and combined effects of ochratoxin A (OA) and Salmonella enterica serovar Gallinarum (S. Gallinarum) on gross and histopathological changes in broiler chickens. One hundred and seventy-six 1-day-old broiler chicks were divided into two groups of 88 chicks each; one group was fed a control mash diet, and the other group was fed a mash diet containing 2 parts/10⁶ OA. On day 14, each group was further subdivided into two groups, with one group inoculated with S. Gallinarum intraperitoneally (1.25×10¹⁰ colony-forming units/0.5ml) whereas the other group was not inoculated with S. Gallinarum. Four birds from each group were sacrificed on 1, 2, 3, 5, 7, 10, 14 and 21 days post inoculation to record pathological changes in different organs. Gross and microscopic changes in OA-fed birds indicated the kidneys and bursa of Fabricius as the primary organs to be affected by this toxin. Gross and microscopic changes due to S. Gallinarum infection indicated the liver and spleen as the primary organs affected by this infection. The effects of OA on the kidney and bursa of Fabricius were enhanced following S. Gallinarum infection. Degenerative changes and interstitial nephritis in the kidneys, and lymphocyte depletion from bursal follicles were more pronounced and were observed earlier in the combination group. In conclusion, data indicate that birds fed OA and infected with S. Gallinarum will demonstrate increased pathology compared with birds fed OA alone or those infected with S. Gallinarum but not fed OA.     

Virulence factors and clonal relationships among Escherichia coli strains isolated from broiler chickens with cellulitis

In this study, we compared Escherichia coli isolates from chickens with avian cellulitis with those from feces of healthy chickens. Cellulitis-derived strains presented phenotypic and genotypic characteristics of greater virulence than did the fecal isolates. Phylogenetic analysis by repetitive extragenic palindromic-PCR showed that, in agreement with their virulence characteristics, the cellulitis isolates form two clonal groups distinct from the fecal isolates.     

Detection of chromosomal blaCTX-M-2 in diverse Escherichia coli isolates from healthy broiler chickens

The rise of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli in food-producing animals is a growing concern for public health. We investigated ESBL producers isolated from broiler chickens in Brazil and characterized 19 CTX-M-2-producing E. coli. The ISCR1 was detected upstream of the chromosome-located gene bla_CTX-M-2, associated with sul-1 type integron structure. CTX-M-2-producing E. coli exhibited different PFGE-types and phylogenetic groups, showing a non-clonal dissemination. The sequence types found (ST93, ST155 and ST2309) have been associated with humans and animals worldwide. Herein, we report the chromosomal location of bla_CTX-M-2 on E. coli, highlighting the risks of multidrug-resistant bacteria in food-producing animals.     

The effects of early exposure of cellulitis-associated Escherichia coli in 1-day-old broiler chickens

Two experiments were performed to test the effect of various field strains of Escherichia coli of cellulitis origin. In the first experiment, 1-day-old broiler chicks were challenged with one of two E. coli field strains using inoculation routes including oral gavage, swabbing of the navel and subcutaneous injection. No cellulitis lesions were produced, although the birds experienced high levels of septicemia/toxemia, characteristic of colibacillosis. The birds that received the E. coli by subcutaneous injection experienced the highest rate of mortality, while those that were challenged by gavage and those that had their navels swabbed experienced lesser rates of mortality. Birds in the second experiment were challenged at 1 day of age with one of three field strains of cellulitis-origin E. coli administered alone or in combination (1:1), which were serially diluted prior to subcutaneous injection. No significant differences in body weight, mortality or cellulitis rates were associated with specific isolates given; however, significant differences were seen with mortality and cellulitis rates according to the dilution of bacteria given. A linear effect was also noted with body weight at 3 weeks, again correlating to the dilution of bacteria that the chicks received.     

Pathology of spleen and liver in turkeys inoculated with Escherichia coli

Lesions of spleen and liver were studied by light and electron microscopy in 3-week-old turkeys inoculated intravenously with Escherichia coli. Spleen and liver of turkeys given avirulent E. coli were hyperaemic and had increased numbers of heterophils. Aggregates of heterophils were in the splenic red pulp and hepatic sinusoids. Turkeys given virulent strains of E. coli had intense heterophil infiltration in spleen and slightly increased numbers of heterophils in liver. In these spleens, reticular sheaths . (ellipsoids) contained large aggregates of heterophils, fibrin, and thrombocytes 2 to 8 hours postinoculation. Turkeys given virulent E. coli and killed in extremis had hypaeremic red pulp and fibrinopurulent exudates and haemorrhage in splenic white pulp. Livers of birds killed in extremis were turgid, friable and engorged with dark blood. Centrolobular hepatocytes were individualised and vacuolated. The dilated sinusoids contained fibrin, erythrocytes, heterophils and bacterial colonies. Phagosomes of hepatic macrophages contained up to 10 structurally normal bacilli, whereas heterophils contained bacilli with altered cell walls. Thrombi, composed of thrombocyte aggregates and fibrin, were common in small hepatic veins. These studies indicate that early bacteraemia with virulent E. coli leads to intense fibrinopurulent splenitis. Progressive bacteraemia is associated with pooling of blood in viscera, intravascular coagulation, and venous thrombosis and bacterial colonisation in liver.     

The contribution of systemic Escherichia coli infection to the early mortalities of commercial broiler chickens

Avian pathogenic Escherichia coli (APEC) are a substantial burden to the global poultry industry. APEC cause a syndromic poultry infection known as colibacillosis, which has been previously associated with broiler chickens over 2 weeks old. We recently reported that the intestinal tract of 1-day-old broilers harbours a rich reservoir of potentially pathogenic E. coli. Prior infections of the reproductive tract of breeders, egg hygiene and transportation all contribute to early colonization of the neonatal gut. Up to one-half of all flock deaths occur in the first week of production, but few data are available describing the contribution of E. coli. In the present study, all dead birds collected on the first daily welfare walk 48 and 72 h after chick placement underwent post-mortem examination. Diseased tissues were selectively cultured for E. coli and isolates subsequently virulotyped using 10 APEC virulence-associated genes (VAGs): astA, iss, irp2, iucD, papC, tsh, vat, cvi, sitA and ibeA. Approximately 70% of birds displayed signs of colibacillosis. Thirty distinct virulence profiles were identified among 157 E. coli. Isolates carried between zero and seven VAGs; ～30% of E. coli isolates carried five to seven VAGs, with 12.7% sharing the same VAG profile (astA, iss, irp2, iucD, tsh, cvi and sitA). Overall, this study demonstrates the significant contribution of E. coli infections to early broiler mortalities. The identification of a diverse E. coli population is unsurprising based on our previous findings. This work emphasizes the need for an effective vaccination programme and provides preliminary data for vaccine production.     

The association of various isolates of Escherichia coli from the United States with induced cellulitis and colibacillosis in young broiler chickens

An experiment was conducted to observe the effects of 10 different avian Escherichia coli isolates in 3-day post-hatch broiler chicks after subcutaneous administration. Isolates were originally obtained from various avian sources throughout the US. Chicks were injected subcutaneously on the ventral surface and necropsied at 7-day intervals for 3 weeks. Cellulitis was produced in all treatments receiving E. coli of cellulitis origin, with the highest incidence occurring 2 weeks post-infection in birds that received an isolate recovered in a previous challenge experiment. Cellulitis was also observed at week 1 post-infection in a small percentage of the birds in two of the treatments receiving E. coli of enteric origin, although lesions disappeared from the group after week 1 post-infection. Septicaemia was the most frequent sequel to challenge and occurred regardless of which isolate was injected. Chicks exposed to cellulitis origin isolates developed septicaemia more frequently than birds challenged with E. coli of non-cellulitis origin. The data implies that cellulitis is unlikely to occur early in the bird's life, since young birds exposed to E. coli frequently develop septicaemia.     

Escherichia coli cellulitis in broiler chickens: clonal relationships among strains and analysis of virulence-associated factors of isolates from diseased birds.

Thirty-nine Escherichia coli isolates from broiler chickens with cellulitis were serotyped and analyzed for clonal relationships by multilocus enzyme electrophoresis. The isolates were further characterized with respect to hemagglutination (HA); serum resistance; antibiotic susceptibility; production of aerobactin, colicin V, and hemolysin; expression of K1 or K5 capsule; sensitivity to cloacin DF13 after treatment with diphenylamine; expression of iron-regulated outer membrane proteins; and virulence in 1-day-old chickens. In addition, the isolates were examined for the presence of DNA sequences related to F1A (fim) and P (pap) fimbriae, aerobactin synthesis (iuc) and transport (iut), hemolysin operon hly, and TraT lipoprotein-induced serum resistance (traT). Only 38.4% of the isolates were typeable with standard O antisera, and of these, serogroups O25 and O78 were the most frequently observed. Multilocus enzyme electrophoresis, based on 20 enzymes, resolved 17 electrophoretic types, forming seven clusters. Isolates from four of these clusters fell into E. coli clone complexes that have been previously reported to be commonly associated with avian colibacillosis. All isolates expressed two to five iron-regulated outer membrane proteins, were resistant to serum and cloacin DF13, and possessed DNA sequences homologous to fim and iuc/iut. Most isolates (72%) were positive for traT, and a majority produced colicin V and aerobactin (92 and 82%, respectively). Assays for the presence of fim and pap DNA sequences, for HA, and for virulence gave variable results but suggest that cellulitis isolates may express F1A and/or other mannose-resistant HA fimbriae different from P and may be virulent in 1-day-old chickens. Our results support the hypothesis that cellulitis in broilers in many cases is caused by E. coli clones identical to other pathogenic avian E. coli strains. Certain clones may be specific to cellulitis, because 25% of the isolates tested belong to clusters not related to known clone complexes.     

Passage of Salmonella through the crop and gizzard of broiler chickens fed with fermented liquid feed

In vivo experiments were conducted in order to investigate the passage and bacterial reduction of Salmonella in the crop and gizzard of chickens when fed two different feeds. The chickens were fed dry conventional feed and fermented liquid feed. The fermented feed contains a relatively high concentration of lactic and acetic acid and lactobacilli. One and three week old broiler chickens were necropsied at short intervals after inoculation with Salmonella Enteritidis. Counts of Salmonella from the crop, gizzard, duodenum, caecum and colon/rectum were obtained. This revealed a sharper decrease of Salmonella in the anterior parts of the gastro-intestinal tract in chickens fed with fermented feed than in chickens fed dry feed. It is therefore concluded that fermented feed improves the barrier formed by the crop and gizzard. The reduction of Salmonella is fully realised in the crop and gizzard. The lower intestinal compartment did not show a substantial effect on the reduction of Salmonella. The performed in vivo method appeared to be an appropriate way to study intervention strategies that aim to control Salmonella by improving the barrier function of the upper gastro-intestinal tract.     

Variations of clinical biochemical parameters of laying hens and broiler chickens fed aflatoxin-containing feed

Two groups of 32 laying hens (Hyssex Brown) and two groups of 32 23-day-old (Hybro) broiler chickens were fed 2.5 and 5 parts/10(6) of aflatoxin in their diet for 4, 8, 16 and 32 days; 16 hens and 32 chicks were maintained as control groups (0 parts/10(6)). After the intoxication period, a clearance period was established of 1, 2, 4 and 8 days. Relative weights of liver and kidneys significantly increased in intoxicated hens, but not in broiler chickens. Histological lesions in both types of bird consisted of hepatic cell vacuolation with fatty infiltration. There was a significant decrease (P< 0.001) in egg production in the 5 parts/10(6) group, which started to recover during the clearance period. No morbidity or mortality due to the aflatoxicosis were observed in either type of bird. In intoxicated laying hens, cholesterol levels were not significantly (P> 0.05) different from control values, but triglyceride levels decreased (P< 0.001) in both intoxicated groups. The effect of aflatoxin on calcium and phosphorus levels was important, because on the 4th day their values decreased significantly. Aspartate aminotransferase (AST) serum levels remained normal, whereas alanino aminotransferase (ALT) activity decreased in both intoxicated groups. The activity of serum lactic dehydrogenase (LDH) and gammaglutamil transferase (GGT) increased significantly. In intoxicated broiler chickens, aflatoxins did not alter (P> 0.05) the biochemical parameters studied, except that the serum calcium concentration was lower in the 5 parts/10(6) group. These data indicated that in intoxicated laving hens, a severe clinical biochemical alteration was produced, and that this together with the hepatic lesions observed in hens and broilers may aid disease diagnosis.     

Haematological and some biochemical changes in experimental fowl typhoid infection in broiler chickens

A study was conducted to evaluate the effect of Salmonella Gallinarum (9, 12) infection on broiler chicks infected through oral and intraperitoneal routes. One hundred and ten broiler chicks (1 week of age) were divided into three groups: group CR (30 chicks) were kept uninfected and served as control group, group OR (40 chicks) were inoculated orally with Salmonella Gallinarum (10⁹ organisms/ml) and group IP (40 chicks) were infected intraperitoneally with Salmonella Gallinarum (10⁹ organisms/ml). Three birds from each group (including those that die) were sacrificed at 3, 5, 7, 14, 21, 28, 35 and 42 days post infection (DPI). Clinical signs were noticed as early as 12 h after intraperitoneal infection and on 3 DPI after oral infection. Higher mortality of 47.50% was observed in intraperitoneally infected chicks as compared to 22.50% in orally infected birds. Decreased body weight and reduced feed intake were observed in chicks in both the infected groups; however, the effects were more marked in chicks infected intraperitoneally. Mean values of haemoglobin, packed cell volume and total erythrocyte count were significantly lower (P?<?0.05) in both orally infected as well as intraperitoneally infected chicks. Leucocytosis observed in both the infected groups was due to increase in the number of heterophils and lymphocytes. Significant decrease (P?<?0.05) in serum total protein and serum albumin was observed in intraperitoneally infected birds only. However, the values of aspartate transaminase and alanine transaminase were increased significantly (P?<?0.05) in both the orally infected and intraperitoneally infected birds.     

Serum biochemical profile and performance of broiler chickens fed diets containing essential oils and pepper

Serum biochemical parameters and performance data were evaluated in broilers fed diets supplemented with antibiotics or essential oils from oregano, sage, rosemary, and pepper crude extract (OLES). Animals (n = 910) were distributed into five treatment groups, with seven replicates of 26 birds in each group: the control group (diet without additives); the group receiving an antibiotic growth promoter diet; and the groups T₅₀, T₁₀₀, and T₁₅₀ (feed supplemented with 50, 100, and 150 mg/kg of OLES, respectively). After 42 days, 55 animals were randomly selected for serum biochemical profile analysis involving pancreatic, renal, and hepatic functions (lipase, amylase, urea, uric acid, aspartate aminotransferase, gamma glutamyltransferase, total cholesterol, high-density lipoprotein, triglycerides, total protein, albumin, globulins, and albumin/globulins ratio). Growth performance of broilers (body weight, weight gain, feed intake, alimentary conversion, and mortality) was also evaluated throughout the experiment period. The increase in serum levels of lipase, uric acid, urea, and aspartate aminotransferase suggests that OLES may cause kidney and liver impairment, mainly, at the higher dose. The OLES dose of 100 mg/kg is suitable for a final body weight and a weight gain similar to those observed in broilers supplemented with antibiotic growth promoters.     

The influence of Escherichia coli strains from different sources and the age of broiler chickens on the development of cellulitis

In two experiments, broilers were challenged with one of several field strains of Escherichia coli to determine whether the source of the E. coli and age of the bird at time of inoculation affected the development of cellulitis lesions. In the first experiment, birds inoculated at 52 days of age with E. coli of faecal, airsacculitis and cellulitis origin exhibited a cellulitis lesion incidence of 47.5, 25 and 77.5%, respectively. This study confirms earlier observations that E. coli strains isolated from cellulitis lesions express a higher propensity for producing these same lesions than other strains, including those associated with airsacculitis. In the second experiment, birds were inoculated at 4, 7, 10, 16, 28, and 52 days of age with an E. coli strain of cellulitis origin and necropsied 2 days post-infection. The resulting incidence of cellulitis ranged from 20% (day 7) to 95% (days 16 and 28), indicating that cellulitis can develop in any age of bird, although the lesions were frequently associated with other manifestations of colibacillosis (perihepatitis, pericarditis, airsacculitis) in birds challenged from 4 to 16 days of age.     

Systemic Mycoplasma synoviae infection in broiler chickens.

Systemic Mycoplasma synoviae infection in 47-day-old broiler chickens with septicaemic lesions and increased carcass condemnation rate is reported. The clinical history included respiratory signs and an enlarged keel bursa. Condemnations at the processing plant were due to airsacculitis and keel bursitis. Involvement of several organs, including the keel bursa, liver, spleen, brain, choroid of the eye, nerves and skeletal muscle associated with vasculitis, and the isolation of M. synoviae from the liver and keel bursa are only occasionally seen in field cases. Random amplified polymorphic DNA analysis of the M. synoviae isolated from the broiler chickens in this study had a different pattern when compared with the reference M. synoviae strains, WVU-1853, MS-H and F10-2AS, and another M. synoviae isolated from broiler breeders from the same company, but had a similar DNA pattern to an M. synoviae isolated from broiler chickens and turkeys owned by the same company. This finding suggests a horizontally acquired infection rather than vertical transmission.     

Phenotypic and genotypic changes in Salmonella enterica subsp. enterica serotype typhimurium during passage in intestines of broiler chickens fed on diets that included ionophore anticoccidial supplements

The effect of continuous in-feed administration of anticoccidial agents on antimicrobial sensitivity and the level of bacterial shedding in poultry experimentally infected with Salmonella enterica subsp. enterica serotype Typhimurium definitive type 104 (DT104) were investigated. On day 0, 1,200 1-day-old Salmonella-free broiler chicks were placed into 50 pens, and the pens were randomly allocated to one of five treatments: nonsupplemented (negative control; T1), monensin at 120 mg/kg of diet (T2), salinomycin at 60 mg/kg of diet (T3), semduramicin at 20 mg/kg of diet (T4), or semduramicin at 25 mg/kg of diet (T5). Each bird was inoculated with a well-characterized strain of serotype Typhimurium DT104 on day 10. On day 49, the birds were euthanatized humanely. Bulk fecal samples were collected on days 13, 43, and 48 and were examined for organisms which had acquired resistance. The genetic basis of acquired resistance was determined from representative samples of isolates. Of 784 Salmonella-selective plates supplemented with antimicrobial agents, only 33 showed growth. These isolates came from all treatment regimens, including the nonsupplemented control. A number of phenotypic changes were observed; these included changes in motility, phage type, and agglutination properties. Supplementation of the diet with an anticoccidial drug does not appear to affect antimicrobial resistance or the level of excretion of salmonellae. Most of the changes observed do not seem to be related to the presence of a supplement in feed. Salmonellae appear to be capable of acquiring antimicrobial resistance and phenotypic changes independently of specific antimicrobial selection pressures.