ABSTRACT Salmonella Heidelberg is an emerging pathogen in Brazilian poultry production. The traditional methods (quicklime, windrowing and tarpaulin-on-surface) used for disinfecting reused poultry litter between flocks does not guarantee its elimination, thus allowing the transmission of this agent from one flock to another. The new tarpaulin-on-surface method with controlled injection of ammonia gas has proven to be effective in its control, however, it is still unknown what dose of ammonia gas is needed to eliminate Salmonella Heidelberg in reused poultry litter. The objective of this study was to evaluate the effect of ammonia gas at different concentrations in sterile poultry litter artificially contaminated with Salmonella Heidelberg. Then, ammonia gas was injected in concentrations of 0.25%, 0.5%, and 1%, and 48 hours later, a sample was collected from each repetition in an entirely randomized design, and bacterial isolation was performed. All treatments, including positive and negative controls, were tested in quadruplicate and the parameters temperature, humidity, pH and water activity were evaluated. In the 0.5% and 1% treated samples the pathogen was not isolated, while in the 0.25% concentration one of the four samples tested was positive. The study reveals that ammonia gas is efficient in killing Salmonella Heidelberg in poultry litter at concentrations of 0.5 % or more within a 48-hour period and that the litter treated with ammonia gas increases its pH and water activity.

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 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 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 The routine use of antimicrobials in animal production for the treatment of infections, disease prevention, or as growth promoters is a predisposing factor for the development and dissemination of antimicrobial resistance. In food industries, sanitizers are used for the control of microbial colonization, and their efficacy depends on contact time and on the dilution of the products used. The present study assessed the effect of 12 antimicrobials and four commercial sanitizers on 18 Salmonella spp. strains isolated from poultry processing plants. None of the evaluated antimicrobials was 100% effective against the tested Salmonella spp. strains; however, 94% of the isolates were susceptible to ciprofloxacin, 77% to amoxicillin + clavulanic acid and to ampicillin, and 72% to enrofloxacin, whereas 100% of the isolates were resistant to penicillin G, 16% to tetracycline, and 11% to sulfonamide. The tested Salmonella spp. strains were 100% inhibited by peracetic acid after five minutes of contact, 0.5% by quaternary ammonium after 15 minutes, and 85.7% by chlorhexidine after 15 minutes. The results indicate the importance of testing of efficacy of antimicrobials used in animal production and in public health to monitor their action and the development of resistance.

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.

Salmonella is traditionally identified by conventional microbiological tests, but the enumeration of this bacterium is not used on a routine basis. Methods such as the most probable number (MPN), which utilize an array of multiple tubes, are time-consuming and expensive, whereas miniaturized most probable number (mMPN) methods, which use microplates, can be adapted for the enumeration of bacteria, saving up time and materials. The aim of the present paper is to assess two mMPN methods for the enumeration of Salmonella sp in artificially-contaminated chicken meat samples. Microplates containing 24 wells (method A) and 96 wells (method B), both with peptone water as pre-enrichment medium and modified semi-solid Rappaport-Vassiliadis (MSRV) as selective enrichment medium, were used. The meat matrix consisted of 25g of autoclaved ground chicken breast contaminated with dilutions of up to 10(6) of Salmonella Typhimurium (ST) and Escherichia coli (EC). In method A, the dilution 10-5 of Salmonella Typhimurium corresponded to >57 MPN/mL and the dilution 10-6 was equal to 30 MPN/mL. There was a correlation between the counts used for the artificial contamination of the samples and those recovered by mMPN, indicating that the method A was sensitive for the enumeration of different levels of contamination of the meat matrix. In method B, there was no correlation between the inoculated dilutions and the mMPN results.

This study was conducted to determine the interference of Salmonella typhimurium lipopolysaccharide (sLPS) on the performance, biological parameters, and histological evaluations of 198 one-day-old male broiler chickens divided into three treatments according to sLPS dose (0, 250, or 500 µg/application/bird) that was applied to the birds every other day, from 15 to 27 days of age. At the end of the experiment (28 days), significant effects were observed on body weight (R= -0.17 and P=0.05), total cholesterol serum levels(R=0.43 and p<0.01), phosphorus (R=0.53 and P<0.01), uric acid (R= -0.38 and P<0.01), C-reactive protein (R=0.68 and p<0.01), serum activity of aspartate aminotransferase (R=0.39 and p<0.01) and alkaline phosphatase (R= -0.39 and p<0.01). According to these results, sLPS mainly affect broiler biological parameters, but also their live performance.

Salmonellosis is an important disease with economic impact as it may affect animal performance and may result in foodborne disease in humans through the eggs and carcass contamination. Regarding the Salmonella control, it is possible to decrease its fecal excretion and the contamination of chicken carcasses by adding organic acids to the feed or drinking water at appropriate times. The aim of this study was to test a blend of organic acids and essential oils in broilers challenged with Salmonella Enteritidis (SE), and to verify the fecal excretion of Salmonella. Sixty broilers were placed in four groups. One group was the negative control. Another group was orally inoculated at 1 day-old with 10(5) CFU/mL of SE as a positive SE control. Two groups (T3 and T4) were orally inoculated at 1 day-old with 10(5) CFU/mL of SE and their feed was separately treated with 0.5 and 1% of organic acids and essential oils, respectively. To assess the fecal excretion of SE, cloacal swabs were collected from all birds at 2, 6, 13 and 20 days after inoculation. The T3 and T4 groups showed a reduction in fecal excretion of SE at 6 and 20 days after inoculation.

The threat to public health represented by Salmonella is at least partially a consequence of its ecology in poultry hosts. Good manufacturing practices in the processing plant can reduce the contamination of poultry products, and critical control point principles are essential throughout the chain production. One procedure adopted in critical points control to prevent and to reduce Salmonella in farms and poultry products is the use of disinfectants. This study aimed at evaluating disinfectant efficiency against Salmonella enteritidis samples isolated from broiler carcasses in Rio Grande do Sul State between 1995 and 1996. The tested disinfectants were: phenol 1:256, quaternary ammonium 1:2500, glutaraldehyde 1:200, and iodine 1:500, with contact times of 5, 10, 15, and 20 in an in vitro test. .Phenolic compounds showed better results, iodine and glutaraldehyde showed intermediary results, and quaternary ammonium presented efficiency at all contact times evaluated in the in vitro test.

Campylobacter was investigated in cecal droppings, feces, and cloacal swabs of 22 flocks of 3 to 5 week-old broilers. Risk factors and the likelihood of the presence of this agent in these flocks were determined. Management practices, such as cleaning and disinfection, feeding, drinkers, and litter treatments, were assessed. Results were evaluated using Odds Ratio (OR) test, and their significance was tested by Fisher's test (p<0.05). A Campylobacter prevalence of 81.8% was found in the broiler flocks (18/22), and within positive flocks, it varied between 85 and 100%. Campylobacter incidence among sample types was homogenous, being 81.8% in cecal droppings, 80.9% in feces, and 80.4% in cloacal swabs (230). Flocks fed by automatic feeding systems presented higher incidence of Campylobacter as compared to those fed by tube feeders. Litter was reused in 63.6% of the farm, and, despite the lack of statistical significance, there was higher likelihood of Campylobacter incidence when litter was reused. Foot bath was not used in 45.5% of the flocks, whereas the use of foot bath associated to deficient lime management increased the number of positive flocks, although with no statiscal significance. The evaluated parameters were not significantly associated with Campylobacter colonization in the assessed broiler flocks.

Dirt floors are used on most Brazilian poultry farms since the construction of concrete floors is very expensive. In vitro tests carried out to verify the effectiveness of disinfectants do not consider the adverse conditions found in poultry farms. Therefore, the present study aimed at evaluating the effect of six commercial disinfectants on the reduction of total and fecal coliforms on the dirt floor of breeder houses. The amount of disinfectant solution to be used per square meter was defined by counting total and fecal coliforms at different soil depths and by analyzing soil physical properties. Coliforms were detected at 0.5 cm, and one liter of disinfectant solution was sufficient for soil saturation at this depth. After that, the efficacy of six commercial products (caustic soda, hydrated lime, phenols 1 and 2, iodine, glutaraldehyde, and quaternary ammonium) in reducing the number of coliforms, after six hours of contact with the dirt floor, was assessed using the most probable number (MPN) method. Escherichia coli specimens isolated from the dirt floor were used to evaluate in vitro effectiveness of disinfectants. Products that yielded the best results in the MPN method were also effective in the in vitro tests. Among the tested disinfectants, hydrated lime was the most efficient, reducing the initial contamination by 2.9 log after six hours of contact with the dirt floor.

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.

Infectious Bursal Disease (IBD) is a chicken disease economically important for the poultry industry in function of the immune depression that it causes. Disease control is made with different vaccines and vaccination programs. In present work, the pathogenicity of 3 intermediate vaccines (I1, I2 and I3), 2 intermediate more pathogenic (IP1 and IP2) and 3 vaccines containing strong virus (F1, F2 and F3) was evaluated. Birds vaccinated with IP1, IP2, F1, F2 and F3 showed significantly lower bursa size in relation to control animals and animals vaccinated with I1, I2 and I3. On the other hand, vaccines I1 and I3 induced antibody titers higher than the control and lower than I2, IP1, IP2, F1, F2 and F3. Histological scores showed that vaccines I1, I2 and I3 induced similar injury degree, although I2 and I3 were not different from the control, whereas I1 was slightly different. Strong vaccines induced more pronounced lesions than the other tested vaccines. These findings suggest that strong vaccines are able to cause severe bursal injuries. However, bursometry and relative weight of the bursa of Fabricius were considered inadequate to evaluate vaccine pathogenicity. Moreover, strong vaccines induced higher antibody titers than the other vaccines, although some intermediate vaccines induced similar titers.