Claudete Serrano Astolfi Ferreira

Prevention of Salmonella Typhimurium colonization and organ invasion by combination treatment in broiler chicks

The effects in broiler chicks of treatment with a competitive exclusion (CE) product, an experimental dietary probiotic, and the abiotic beta-glucan on cecal colonization, organ invasion, and serum and intestinal IgG and IgA levels to Salmonella challenge was evaluated. Four groups of 1-d-old chicks were treated by oral gavage on d 1 with an appropriate dose of a commercial CE product. Three groups received daily doses of probiotic, beta-glucan, or both, for 6 d. Three other groups were fed daily from d 1 onwards with probiotic, beta-glucan, or both. Subgroups of 30 chicks from each group were challenged on d 1, 9, 16, or 23 with 10(7) cfu/ mL of Salmonella Typhimurium (1769NR) and killed 7 d later. Control groups were maintained untreated and remained unchallenged (negative control), or were challenged with Salmonella Typhimurium (1769NR; positive control), as described above. Cecum, liver, and spleen samples were examined for the presence of Salmonella, whereas serum and intestinal fluid samples were assayed for total antibody (IgG and IgA) concentrations. Data were analyzed by 1-way ANOVA, and means were compared using Duncans multiple range test. In comparison with other treatments, those involving CE product and beta-glucan, with or without probiotic during the first week, resulted in a superior inhibition of cecal colonization and organ invasion by Salmonella and also offered a higher level of protection (P < 0.05). During the second week, treatments containing experimental dietary probiotic and beta-glucan, with or without CE product, resulted in an inhibition of liver invasion (P < 0.05). The IgA levels were significantly higher (P < 0.05) in intestinal fluid compared with serum, whereas IgG had low levels. The results in the first and third week indicate that combination treatments involving CE product, probiotic, and beta-glucan are a more effective control of Salmonella colonization than the corresponding individual preparations.

Virulence properties of Escherichia coli isolated from ostriches with respiratory disease.

Eight Escherichia coli isolates from ostriches with respiratory disease were investigated for the presence of genes encoding the following adhesins: type 1 pili (fim), pili associated with pyelonephritis (pap), S fimbriae (sfa), afimbrial adhesin (afaI), temperature regulated adhesin, curli (crl, csgA) and temperature-sensitive hemagglutinin (tsh). Genes for heat labile (LT) and heat stable (STa and STb) enterotoxins, Shiga toxins (stx1 and stx2), cytotoxic necrotizing factor 1 (cnf), alpha-haemolysin (hly) and aerobactin (aer) production were also investigated. Other characteristics investigated were the presence of hemagglutination activity, growth on an iron-deficient medium, aerobactin production, serum resistance, adherence to chicken tracheal cells, pathogenicity for day-old chicks, and serogroup. Serogrouping showed that four isolates belonged to serogroup O2, two to serogroup O78, one to serogroup O9, and one to serogroup O21. The virulence genes found were: fim in all eight isolates, csgA in seven, aer in six, and pap, crl and tsh in one isolate each. All isolates analyzed were positive for mannose-resistant hemagglutination, adhered in vitro to ciliated tracheal epithelium, grew on iron-deficient medium, and showed serum resistance. Pathogenicity tests on day-old chickens revealed one highly pathogenic isolate, three of low pathogenicity and four isolates with intermediate pathogenicity.

Distribution of virulence genes sefC, pefA and spvC in Salmonella Enteritidis phage type 4 strains isolated in Brazil

The distribution of virulence genes, sefC, pefA and spvC, was investigated in 110 Salmonella Enteritidis phage type 4 strains by polymerase chain reaction. Their influence in the caecal colonization and invasion of liver and spleen of one-day-old chickens was studied. Eight isolates were negative for the spvC gene, three for the pefA gene and one, for the sefC gene. These results allowed grouping the strains into four genotypes. Presence of these genes did not influence bacteria invasion in the liver and spleen of the chickens ten days after infection, although the presence of more than one fimbrial gene can be related to caecal colonization.

Comparison of experimental competitive-exclusion cultures for controlling Salmonella colonization in broiler chicks

Esse estudo avaliou a eficacia de tres diferentes tipos de culturas de exclusao competitiva (EC-A, EC-B e EC-C) contra Salmonella Kedougou (SK), amostra NCTC 12173, resistente ao acido Nalidixico (NalR), em pintos de um dia de idade, utilizando-se 4 tratamentos, em tres repeticoes. A media logaritmica de Salmonella por gramas de fezes foi de 0,41 para o grupo tratado com a EC-A, contendo uma mistura de culturas bacterianas aerobias e anaerobias, derivada de conteudo cecal de uma ave adulta; 1,22 no grupo tratado com a EC-B, contendo culturas bacterianas aerobias; 1,00 no grupo tratado com a EC-C, contendo culturas bacterianas anaerobias e 6,64 no grupo controle positivo. A porcentagem de aves colonizadas variou de 10 a 23,33% nos grupos tratados e foi de 63,33% no grupo controle. Uma boa protecao (76,66 a 90%) foi obtida em todos os tratamentos, sendo que a menor protecao foi verificada com os produtos experimentais contendo somente culturas de bacterias aerobias ou anaerobias. Os resultados sugerem que o uso de misturas de culturas aerobias e anaerobias pode ser efetivo para o uso na exclusao competitiva contra SK em frangos de corte.

Virulence and molecular aspects of Bordetella avium isolated from cockatiel chicks (Nymphicus hollandicus) in Brazil

Bordetella avium is an opportunistic pathogen that presents tropism for ciliated epithelia, leading to upper respiratory tract disease in turkeys. This agent has also been associated with Lockjaw Syndrome in psittacine birds, but literatures describing the importance of this agent in such species are rare. The purpose of the present study was to report the first outbreak of B. avium infection in juvenile cockatiels demonstrating the Lockjaw Syndrome in Brazil and to investigate the antimicrobial resistance profile and phenotypic and genotypic characteristics of these strains. Surprising, the strains obtained from five infected cockatiel chicks from three different breeders from different Brazilian states showed a clonal relationship using the Pulsed Field Gel Electrophoresis and Single Enzyme Amplified Fragment Length Polymorphism techniques. The virulence potentials of the B. avium strains were assessed using tracheal adherence and cytotoxic effects on a VERO cell monolayer.

Hemagglutinating properties of Salmonella enterica serovar Enteritidis isolated from different sources

Twenty-five strains of Salmonella enterica serovar Enteritidis isolated from different sources were examined for hemagglutinating activity. Bacteria cultured in different media induced hemagglutination of human erythrocytes, but no reaction was observed with erythrocytes from other animal species. The hemagglutinating expression activity was better for cultures on CFA agar at 37oC than other conditions examined. The hemagglutination was inhibited by D-mannose, D-mannitol, melibiose, D-raffinose, L-rhamnose and sucrose. The absence of cell-surface appendages in electron microscope examinations suggested a nonfimbrial hemagglutinin. The data suggest that Salmonella Enteritidis produces nonfimbrial mannose-sensitive hemagglutinin, specific for human erythrocytes, which could be extracted in soluble form.

Analysis of the Virulence of an Atypical Enteropathogenic Escherichia coli Strain In Vitro and In Vivo and the Influence of Type Three Secretion System

Atypical enteropathogenic Escherichia coli (aEPEC) inject various effectors into intestinal cells through a type three secretion system (T3SS), causing attaching and effacing (A/E) lesions. We investigated the role of T3SS in the ability of the aEPEC 1711-4 strain to interact with enterocytes in vitro (Caco-2 cells) and in vivo (rabbit ileal loops) and to translocate the rat intestinal mucosa in vivo. A T3SS isogenic mutant strain was constructed, which showed marked reduction in the ability to associate and invade but not to persist inside Caco-2 cells. After rabbit infection, only aEPEC 1711-4 was detected inside enterocytes at 8 and 24 hours pointing to a T3SS-dependent invasive potential in vivo. In contrast to aEPEC 1711-4, the T3SS-deficient strain no longer produced A/E lesions or induced macrophage infiltration. We also demonstrated that the ability of aEPEC 1711-4 to translocate through mesenteric lymph nodes to spleen and liver in a rat model depends on a functional T3SS, since a decreased number of T3SS mutant bacteria were recovered from extraintestinal sites. These findings indicate that the full virulence potential of aEPEC 1711-4 depends on a functional T3SS, which contributes to efficient adhesion/invasion in vitro and in vivo and to bacterial translocation to extraintestinal sites.

Lethal Factor to Mice Produced by Escherichia coli Isolated from Chickens with Swollen Head Syndrome

A lethal toxin similar to Bacillus cererus lethal toxin was detected in the culture supernatants of Escherichia coli isolated from chickens with swollen head syndrome. The lethal activity was heat‐labile, protease‐sensitive and killed mice within 10 min. The light microscopy of the histopathological studies revealed that the principal organ affected by this toxin was the lung but the liver and kidneys also showed lesions. The relevance of this lethal activity from E. coli remains to be determined.

Isolation of avian nephritis virus from chickens showing enteric disorders

Abstract Runting-stunting syndrome (RSS) is one of the diseases associated with many detected viruses. In Brazil, there were reports of several enteric disease outbreaks in chickens in which avian nephritis virus (ANV) was detected; however, the role of ANV in the outbreaks and whether the virus was a causative agent of these cases of enteric diseases were not determined. The aim of this study was to isolate ANV in specific pathogen-free (SPF) chicken embryonated eggs (CEE) from the enteric contents of chickens showing signs of RSS. For this purpose, 22 samples of chicken enteric contents that were positive only for ANV were inoculated into 7 and 14-day-old SPF-CEE via the yolk sac route and incubated for 5 d, with a total of 3 passages. Virus isolation was confirmed by the presence of embryo injuries, detection of viral RNA by RT-PCR, and visualization of viral particles using electron microscopy. Therefore, the 7-day-old inoculated embryos showed dwarfism, gelatinous consistency, hemorrhage, and edema in the embryos, whereas the 14-day-old did not show any alteration. Viral RNA was detected in the embryos of both ages of inoculation, and the same viral particles were visualized. The embryos from the mock group showed no alteration and were negative for all the tests. The viral cDNA was sequenced, and the molecular and phylogenetic analyses showed that the Brazilian isolates are more related with the ANV-1 serotype group; the sequences of these isolates showed a high percentage of nucleotide (86.4 to 94.9%) and amino acid (92.3 to 98.7%) similarity with other sequences from China, Japan, Australia, and the United States that belong to this serotype previously classified group. In this study, we isolated 8 samples of ANV in SPF-CEE from enteric content samples from chickens with RSS. In doing so, we showed the pathological injuries to the embryo caused by the virus and the molecular characterization of a part of the ORF 1b gene of the virus.