ABSTRACT In recent years, egg production has had an intense growth in Brazil, and Brazilian egg consumption per capita has significantly increased in the last decade. To reduce sanitary and financial risks, decisions regarding the production and health status of the flock must be made based on objective criteria. Our aim was to determine the main “input” variables for the prediction of egg production performance in commercial laying breeder flocks using an ANN model. The software NeuroShellClassifier and NeuroShell Predictor were used to build the ANN. A total of 26 egg-production traits were selected as input variables and eight as output variables. A database of 44,120 Excel cells was generated. For the training and validation of the models, 74.9% and 25.1% of the data were used, respectively. The accuracy of the ANN models was calculated and compared using the analysis of coefficient of multiple determination (R2), mean squared error (MSE), and an assessment of uniform scatter in the residual plots. The models for the outputs “weekly egg production,” “weekly incubated egg,”, “accumulated commercial egg,” and “viability” showed an R2 greater than 0.8. Other models yielded R2 values lower than 0.8. The ANN predicts adequately eight egg-production traits in the breeders of commercial laying hens. The method is an option for data management analysis in the egg industry, providing estimates of the relative contribution of each input variable to the outputs.

ABSTRACT Due to the genetic similarity of pathogenic Escherichia coli isolated from birds and pathotypes of human origin, it is suggested that they have a common ancestor and may exchange virulence-associated genes. This study aimed to detect virulence-associated genes in E. coli strains isolated from the Red-browed Amazon parrot (Amazona rhodocorytha) kept at a conservation institute in Brazil. High genetic variability in virulence was observed, since 12 virulence profiles were found among 14 strains. The number of virulence-associated genes of single strains ranged from 5 to 22 out of 33 genes tested, and only one strain did not present any virulence genes. Regarding adhesion genes, most strains presented from two to five genes, and crlA (85.7%) and fimC (85.7%) were the most frequent. Frequencies were similar for invasion and iron acquisition genes. Variations among genes were observed for serum resistance and toxin-related genes. Some of the E. coli strains isolated from parrots presented virulence genes that are commonly associated with pathotypes of human origin, including newborn meningitis E. coli, uropathogenic E. coli, and sepsis-associated E. coli. It is noteworthy that some of these genes were present in the majority of the analyzed strains. Our results indicate that these strains detected in clinically healthy parrots can be potential reservoirs of several virulence-associated genes. These genes can be transmitted to other E. coli strains, including those that affect humans. These E. coli strains present a high pathogenic potential of virulence-associated genes in extraintestinal pathogenic E. coli strains.

ABSTRACT Brazil is the largest exporter of chicken meat and poultry farming is one of the most important productive segments, despite major losses due to the bacterium Escherichia coli, which is also a zoonotic microorganism. The objetive of this study was to isolate E. coli and to evaluate its transmissibility potential from the field to chicken meat using the Pulsed Field Gel Electrophoresis (PFGE) technique. Environmental samples (poultry litter, soil and water) were collected from broiler farms located in the South of Brazil where the majority of the Brazilian poultry production occurs. In addition, chicken meat (gizzard, heart, drumette and tulip) samples were collected from local supermarkets. As results, 47.36% of the samples were positives for E. coli. Furthermore, 10 pairs of clones of E. coli were found always in the same substrate (two water-water pairs; three soil-soil pairs and five meat-meat pairs) using PFGE. These findings suggest that certain strains of E. coli may have habitat preferences, making the transfer from one substrate type to another more difficult to occur. Moreover, since no clones were found between environmental samples and chicken meat, it is possible to imply a low risk of E. coli transmissibility throughout the chicken meat production chain.

ABSTRACT Salmonella spp. remain among the most important agents of foodborne diseases worldwide. The importance of Salmonella spp. in public health is linked to their wide range of antimicrobial resistance and to their pathogenicity and virulence in both human and animal hosts. The aim of this study was to determine the antimicrobial resistance patterns for Salmonella serotypes isolated from poultry sources in Brazil and to detect virulence-associated genes and verify their association with specific serotypes. A total of 163 strains of Salmonella enterica isolated from poultry sources in Southern Brazil were selected, and each belonged to one of 11 different serotypes. They were tested against ten antibiotics and examined for the presence of 26 virulence-associated genes by PCR. S. Typhimurium, S. Bredeney, S. Schwarzengrund and S. Tennessee showed the highest overall resistance rates. Approximately 18% of Salmonella strains were classified as multidrug-resistant strains. Our results indicate associations between antimicrobial resistance and specific serotypes. Most of the investigated genes presented a high frequency and a regular distribution, regardless of the serotype. Eight genes are positively or negatively associated with at least one serotype. The observed associations between antimicrobial resistance and specific serotypes are useful in developing specific control and treatment measures for each serotype. Despite the virulence genes being evenly distributed among the serotypes, some of these genes are associated with specific serotypes, and sefA, sopEand lpfA were selected as possible markers of Salmonella serotypes.

ABSTRACT Antimicrobial resistance is a serious public health problem and Salmonella spp. is highly resistant to antimicrobial agents. Biofilms are important in the food industry due to their formation on products, utensils, and surfaces and the difficulty in their removal. The objective of this study was to assess extended spectrum beta-lactamase (ESBL) production, antimicrobial resistance, and biofilm production of Salmonella isolated from poultry slaughterhouses. Antimicrobial susceptibility was assessed by the disk diffusion assay and ESBL by double diffusion disk assay using the beta-lactamase inhibitor (amoxicillin+clavulanate). The antimicrobials tested were: ampicillin, amoxicillin+clavulanate, aztreonam, ceftazidime, cefotaxime, chloramphenicol, gentamicin, enrofloxacin, sulfonamide, and tetracycline. Serovars Infantis, Panamá, and Tennessee were found to produce ESBL. All serovars were sensitive to tetracycline, and S. Brandenburg was sensitive to all drugs tested. Serovars Panamá, Anatum, Infantis, and Schwarzengrund were moderate biofilm producers at 3 ºC and 9 ºC±1 ºC, respectively, showing possible adaptation of these serovars to these temperatures. Antimicrobials should be used with caution because of the levels of resistance observed and because of ESBL production, and hygiene and sanitary measures should be enhanced to minimize the adhesion of biofilm-forming Salmonella serovars at refrigeration temperatures.

ABSTRACT Fowl cholera is a contagious disease that results from infection by the bacterium Pasteurella multocida. This microorganism is extensively distributed among animal species, but little is known regarding it’s pathogenesis and specificity to various hosts. Many studies using pathogenicity evaluation methods are subjective and difficult to quantify because they are often only involved in the observation of the lethal capacity of the agent in experimental inoculation. Due to a lack of more consistent data, this study aimed to establish a classification model of P. multocida pathogenicity in mice using strains isolated from poultry and swine. A total of 94 strains of P. multocida isolated from clinical cases of FC and from lungs of swine were tested. A volume of 0.1 mL of bacterial suspension was obtained from the concentration of 106 CFU/mL and inoculated by an intraperitoneal route in five mice. The animals were observed every six hours over seven days. In addition to the mortality observed, the time of death and gross lesions were also analyzed. The Pathogenicity Indexes obtained showed significant differences (p<0.05) according to the origin of the strains. Likewise, the number of gross lesions and isolation percentages were also varied (p<0.05) among strains isolated from poultry and swine. From the observed ratios, the isolates were grouped into three pathogenicity classes: high, medium and low. This study proposed a consistent measurement and classification of P. multocida pathogenicity. The obtained results will be used to generate other adjusted models, as well as to form the basis for disease diagnosis.

ABSTRACT The Infectious Bursal Disease (IBD) is a contagious viral disease that affects young chickens and may cause high morbidity and mortality. As the virus is very resistant to the environment, vaccination is required in case of high infection pressure. Due to variations in the virulence degree of the vaccines available to control IBD, this study aimed at evaluating the pathogenicity and immunogenicity of three types of vaccines. In total, 220 one-day-old specific pathogen free (SPF) chickens were immunized with recombinant, immune-complex and intermediate vaccines, or not vaccinated (55 birds per group) and challenged with IBD G11 strain on day 25. On days 25, 30, and 35, the Bursa of Fabricius (BF) were submitted to gross and histological examination, and serum samples were submitted to ELISA to determined anti-IBD antibody titers. On day 23, chickens were submitted to the test of hypersensitivity to phytohemagglutinin to evaluate the immunosuppressive effect of vaccines on the cell-mediated immunity. The results have indicated that the immune-complex vaccine induced the most severe BF lesions, whereas the recombinant vaccine preserved BF tissue and cell integrity. The three evaluated vaccines induced humoral immunity of similar intensity. The cellular reaction to phytohemagglutinin of the chickens immunized with recombinant and immune-complex vaccines was less severe compared with the unvaccinated chickens. In conclusion, these results indicate that the immune-complex vaccine was the most pathogenic and that all vaccines were effective in protecting SPF chickens against IBD.

ABSTRACT Although Pasteurella multocida is a member of the respiratory microbiota, under some circumstances, it is a primary agent of diseases , such as fowl cholera (FC), that cause significant economic losses. Experimental inoculations can be employed to evaluate the pathogenicity of strains, but the results are usually subjective and knowledge on the pathogenesis of this agent is still limited. The objective of this study was to establish a new methodology for classifying the pathogenicity of P. multocida by formulating a standard index. Strains isolated from FC cases and from swine with respiratory problems were selected. One hundred mL of a bacterial culture of each strain, containing 106 CFU, was inoculated in 10 one-day-old broilers. Mortality after inoculation, time of death (TD), and the presence of six macroscopic lesions were evaluated over a period of seven days post-inoculation (dpi). A Pathogenicity Index Per Bird (IPI), ranging 0 to 10, was calculated. Liver and heart fragments were collected to reisolate the bacteria. Blood was collected from the surviving birds, and an ELISA test was carried out to detect specific antibodies. The median of the pathogenicity indices, the number of lesions and the rate of bacteria reisolation were significantly different (p<0.05) among the origins of the isolates (p<0.05). The pathogenicity index developed in this study allows the classification of Pasteurella multocida pathogenicity and may be an alternative to the pathogenicity models currently used for screening.

The ability of Pasteurella multocida to invade and multiply in its host is enhanced by the presence of the capsule, one of the most important virulence factors for this bacterium. Capsular typing methods are often used in epidemiological and pathogenesis studies of this agent. Five different serogroups have been identified based on serological typing. However, such tests are laborious, and agglutination of homologous antiserum may fail. The aim of this study was to develop a multiplex PCR protocol for the identification of the hyaD-hyaC and dcbF genes specific to serogroups A and D, respectively, and to compare these results with those of phenotypic tests for 54 strains isolated from fowl cholera cases in southern Brazil. The kappa coefficient and chisquare statistics were calculated to assess the agreement between the diagnostic methods and to determine the significance of the results, respectively. The multiplex PCR was able to detect the evaluated genes. Forty-nine strains (90.74%) were classified into serogroup A, and only two isolates (3.7%) were not identified as belonging to any of the serogroups analyzed. In contrast, with the phenotypic tests, only 41 strains (75.93%) were classified into serogroup A and 11 samples (20.37%) were unidentifiable. Of the strains analyzed, 70.37% were classified into the same serogroup (A) by both methods, and the kappa coefficient (k = 0.017) indicated poor agreement between the tests. Thus, multiplex PCR is an alternative for P. multocida capsular typing, as it allows the simultaneous and rapid detection of genes and also provides a greater strain-typing capacity.

Broiler chicks belonging to two poultry companies, A and B, with different breeders' vaccination programs were challenged with a very virulent strains of infectious bursal disease virus (vvIBDV), genotyped as G11. Birds were separated in four groups, two vaccinated at the first day of life and two unvaccinated. They were then challenged at the 1st, 4th, 7th, 10th, 13th, 16th, 19th and 22nd days. At every day of challenge, before and after the procedure, the following data were collected from each group: Bursa of Fabricius (BF) relative weight, BF diameter, BF for histologie examination, serum for measuring antibodies against IBDV through the ELISA and clinical evaluation of IBD. The results obtained have shown a non-significant drop in antibody level between the vaccinated and the unvaccinated groups. When analyzing the different results, it could be established that an ELISA titre of 3,4 log10 was the cutoff point between healthy and sick birds. Regression equations were built to determinate the best moment for vaccination and also the ELISA log titre birds what could present in a given age. Based on that, chicks from Company A should receive a vaccine against IBD from the 6th to 7th day of age, while the ones from Company B should get it between the 11th and the 12th day of age. Finally, the overall results suggest that the birds should not be vaccinated at one day old, and also that the breeders' different vaccination schemes resulted in progenies with different levels of maternal protection, and as a consequence the same vaccination plan should not be applied indiscriminately to broilers from different poultry companies.