Depuis la découverte de la pénicilline par Sir Alexander Fleming, les antibiotiques ont joué un rôle primordial et incontestable en médecine moderne en aidant à combattre les infections bactériennes. Cependant, les bactéries ont la capacité de se protéger par différents moyens des molécules antibiotiques. La surutilisation de ces molécules a accéléré le phénomène de résistance aux antibiotiques, rendant difficile, voire impossible, le traitement de certaines maladies infectieuses par cette approche. La résistance aux antibiotiques est une problématique d’envergure mondiale qui touche aussi négativement l’aquaculture, où les infections bactériennes peuvent causer d’importantes pertes économiques. L’une de ces bactéries est Aeromonas salmonicida subsp. salmonicida, l’agent étiologique de la furonculose. Bien qu’il fût déjà connu que plusieurs souches de cette bactérie étaient porteuses de plasmides conférant des résistances aux antibiotiques, l’ampleur de la problématique était encore inconnue. Les bactériophages (phages) sont des virus infectant spécifiquement les bactéries. Cette capacité à lyser les bactéries leur a valu d’être utilisés dans un contexte thérapeutique presque dès leur découverte au début du 20e siècle. Cependant, l’avènement des antibiotiques a fait en sorte que la thérapie par les phages a été oubliée dans plusieurs pays occidentaux. Maintenant que la résistance aux antibiotiques est devenue une inquiétude pour la pérennité de notre société, plusieurs études suggèrent que la thérapie par les phages pourrait être une alternative ou un complément aux traitements par antibiotiques. La présente thèse avait comme objectifs : (1) d’explorer la diversité génomique causant une résistance aux antibiotiques chez A. salmonicida subsp. salmonicida et (2) d’investiguer le potentiel d’un traitement par les phages pour contrer les infections causées par cette bactérie. Il a été possible de mettre à jour et de caractériser cinq nouveaux plasmides avec des gènes de résistance aux antibiotiques chez A. salmonicida subsp. salmonicida. De plus, la présence de deux de ces plasmides (pAB5S9b et pSN254b) causent une résistance à tous les antibiotiques approuvés par le gouvernement canadien pour une utilisation par l’industrie piscicole. Avant d’investiguer la diversité des phages infectant A. salmonicida subsp. salmonicida, il était crucial de mieux connaître la bactérie d’intérêt. Plusieurs phages sont connus pour avoir un spectre lytique étroit, n’infectant ainsi que certaines souches ou certaines sousespèces d’une bactérie. Or, la structure intra-espèce d’A. salmonicida était encore mal définie. De plus, l’une des sous-espèces d’A. salmonicida, pectinolytica, est considérée comme mésophile avec la capacité de croître à 37°C, alors que les autres sous-espèces, comme salmonicida, sont limitées à des températures d’environ 20°C et sont par conséquent qualifiées de psychrophiles. En caractérisant de nouvelles souches mésophiles, mes travaux ont mis en lumière que les séquences d’insertion peuvent être une raison pour expliquer cette dichotomie. De plus, il a été possible de démontrer une grande diversité génétique chez les souches mésophiles, comparativement à celles psychrophiles. Afin de vérifier le potentiel d’un traitement par les phages contre la furonculose, trois phages spécifiques à A. salmonicida subsp. salmonicida ont été isolés de l’environnement. L'ADN de ces phages, en plus de celui de neuf autres disponibles à la collection Félix d’Hérelle, a été séquencé à haut-débit sur un appareil MiSeq d’Illumina. En comparant ces séquences génomiques à celles déjà disponibles publiquement, il a été possible de déterminer six groupes génomiques de phages contre A. salmonicida subsp. salmonicida. Les 12 phages disponibles pour la présente étude ont été testés sur 65 souches d’A. salmonicida (incluant des sous-espèces autres que salmonicida), permettant de dresser un portrait de la capacité lytique de chacun de ces virus. Cette analyse a mis en lumière trois groupes de phages ayant des capacités lytiques variables. De plus, il a été possible de montrer que d’autres sous-espèces d’A. salmonicida psychrophiles peuvent être infectées par les phages isolés à partir de la sous-espèce salmonicida. Cependant, les souches mésophiles d’A. salmonicida sont insensibles à ces phages. Cette étude doctorale a montré que la résistance aux antibiotiques est un problème d’envergure dont l’ampleur était insoupçonnée chez A. salmonicida subsp. salmonicida. Elle a aussi permis d’investiguer le potentiel de la thérapie par les phages. / Since the discovery of penicillin by Sir Alexander Fleming, antibiotics have played a paramount and indisputable role in modern medicine in helping to treat bacterial infections. However, bacteria have the ability to protect themselves against antibiotics by various mechanisms. The overuse of these molecules has accelerated the phenomenon of antibiotic resistance, making it difficult, if not impossible, to treat certain bacterial infections. Antibiotic resistance is a global problem that also negatively affects aquaculture, where bacterial infections can cause significant economic losses. One of these bacteria is Aeromonas salmonicida subsp. salmonicida, the etiologic agent of furunculosis. Although it was already known that several strains of this bacterium were carriers of plasmids conferring resistance to antibiotics, the extent of the problem was still unknown before this study. Bacteriophages (phages) are viruses specifically infecting bacteria. Their ability to lyse bacteria has been used in a therapeutic context almost as soon as they were discovered at the beginning of the 20th century. However, the advent of antibiotics has meant that phage therapy was forgotten in several Western countries. Now that antibiotic resistance has become a significant concern for the sustainability of our society, several studies suggest that phage therapy could be an alternative or supplement to antibiotic treatments. The objectives of this thesis were: (1) to explore the genomic diversity causing resistance to antibiotics in A. salmonicida subsp. salmonicida and (2) to investigate the potential of phage therapy to treat infections caused by this bacterium. Five new plasmids conferring antibiotic resistance to A. salmonicida subsp. salmonicida were discovered and characterized. Two of these plasmids, pAB5S9b and pSN254b, cause resistance to all antibiotics approved by the Canadian government for use in the fish industry. Before investigating the diversity of phages infecting A. salmonicida subsp. salmonicida, it was crucial to better know the bacterium of interest. Several phages are known to have a narrow host spectrum, infecting certain strains or subspecies. Until the present doctoral study, the intra-species structure of A. salmonicida was poorly defined. In addition, one of the subspecies of A. salmonicida, pectinolytica, is considered mesophilic with the ability to grow at 37°C, while other subspecies, such as salmonicida, are limited to growth temperatures around 20°C and are therefore considered psychrophilic. By characterizing new mesophilic strains, we found that insertion sequences may be a reason for this dichotomy. In addition, it was possible to demonstrate a high genetic diversity in mesophilic strains compared to psychrophilic strains. In order to verify the potential of phage treatment against furunculosis, three phages specific to A. salmonicida subsp. salmonicida were isolated from the environment. The genomic DNA of these phages, in addition to that of nine other phages available at the Felix d'Hérelle collection, was sequenced on an Illumina MiSeq device. By comparing these genomic sequences to those already available publicly, it was possible to determine six genomic groups of phages infecting A. salmonicida subsp. salmonicida. The 12 phages available were tested on 65 strains of A. salmonicida (including subspecies other than salmonicida), providing the host range of each virus. This analysis revealed three groups of phages with variable lytic capacities. In addition, it was possible to show that other psychrophilic subspecies of A. salmonicida can be infected by phages isolated from the subspecies salmonicida. However, the mesophilic strains of A. salmonicida are insensitive to these phages. This doctoral study showed that resistance to antibiotics is a large-scale problem in A. salmonicida subsp. salmonicida, and that phage therapy may represent one of the solutions to the growing concern

Eine effektive Strategie zur Verhinderung der Ausbreitung von Infektionskrankheiten in der Aquakultur ist die Anwendung geeigneter Präventionsmaßnahmen, insbesondere die Impfung von Fischen. Das effektivste Impfverfahren stellt die individuelle Injektion des Impfstoffes dar. Bei Milliarden von Fischen jährlich ist dies jedoch sehr zeit- und kostenintensiv. Bei Säugetieren gilt Niederfrequenz-Sonophorese (LFS) als eine der fortschrittlichsten Technologien zur transdermalen Verabreichung von Wirkstoffen. So entstand die Idee, bei Fischen die Aufnahme von Wirkstoffen während einer Tauchbadbehandlung mit Hilfe von LFS zu verbessern. Die vorliegende Arbeit zeigt, dass mit einer Beschallungsintensität von etwa 60 mW/cm^2 die Aufnahme eines Bakterienimpfstoffs in das Kiemengewebe von Regenbogenforellen um den Faktor 240 erhöht werden konnte. Bei dieser Intensität traten geringe oder keine Nebenwirkungen auf. Bei höheren Beschallungsintensitäten wurde eine noch höhere Aufnahme des Bakterienimpfstoffes, aber auch schädliche Nebenwirkungen beobachtet. Darüber hinaus zeigte LFS eine durch eine lokale Entzündungsreaktion und Aktivierung von T-Helferzellen in den Kiemen charakterisierte, Adjuvans-ähnliche Wirkung. Ein Impfversuch mit Koi Karpfen und einem inaktivierten Impfstoff gegen das Koi-Herpesvirus zeigte, dass LFS das Potential hat, den mit einer Tauchbadimpfung erzielbaren Immunschutz zu verbessern. LFS konnte auch für die Verabreichung anderer Substanzen wie Antibiotika verwendet und die Aufnahme von Oxytetracyclin um den Faktor fünf erhöht werden. Zusammenfassend lässt sich sagen, dass die Effektivität einer Tauchbadimpfung von Fischen mit LFS durch die gesteigerte Impfstoffaufnahme und dessen Adjuvans-ähnlichen Wirkung verbessert werden kann. Darüber hinaus könnte mithilfe LFS die therapeutische Dosis von Antibiotika bei Badbehandlungen verringert werden. Es erfordert jedoch noch weitere Studien, um diese Technologie aus dem Labor in die Praxis zu übertragen. / In aquaculture, the use of prevention methods, such as vaccination of fish, is an effective strategy to avoid infectious diseases. However, the most effective route of vaccination for fish is the one-by-one intraperitoneal injection, which that is very laborious and expensive to apply for billions of fish every year. Low Frequency Sonophoresis (LFS) has been recognized as one of the most advanced technologies in transdermal delivery of substances in mammals. Thus, it has been suggested to use LFS to enhance the uptake of substances in fish during bath treatments. The present study shows that a low sonication intensity of ca. 60 mW/cm^2 at 37 kHz increased the uptake of a bacterin into the gill tissue of rainbow trout by up to a factor of 240. At this intensity, no or only minimal side effects occurred. At higher sonication intensities, an even higher bacterin uptake but also deleterious side effects were observed. In addition, LFS showed an adjuvant-like effect characterized by a local inflammatory response and T-helper cell activation in the gills. A vaccination trial with koi carp and an inactivated vaccine against the Koi Herpes Virus (KHV) showed that LFS has the potential to enhance the immune protection achieved by immersion vaccination. In addition LFS can also be used for the administration of other substances, such as antibiotics, here we showed that the uptake of Oxytetracycline could be increased by factor five. In summary, the efficacy of the immersion vaccination of fish could be improved by low-frequency ultrasound due, to the increased vaccine uptake along with its adjuvant-like effect. Furthermore, LFS could also reduce the required therapeutic dose of antibiotics in bath treatments, making them more effective, cheaper and environmentally friendly. However, further practical studies will be required to transfer this technology from the lab to the field.

O copolímero biodegradável poli(3-hidroxibutirato-co-3-hidroxihexanoato) P(3HB-co-3HHx) é um polihidroxialcanoato (PHA) que apresenta várias aplicações. A bactéria Burkholderia sacchari acumula P(3HB-co-2mol%3HHx), a partir de glicose e ácido hexanoico. Com o objetivo de obter P(3HB-co-3HHx) com diferentes teores de 3HHx por B. sacchari, foram construídas linhagens recombinantes, contendo genes do operon phaPCJ de Aeromonas spp. Os recombinantes produziram P(3HB-co-3HHx), a partir de ácidos hexanoico, láurico e linoleico, com teores de 3HHx entre 1,88-18 mol%. Experimentos em biorreator com o recombinante, alimentada na fase de acúmulo por glicose 140 g/L e ácido hexanoico entre 0-45 g/L, resultaram copolímeros com composições variando de 0 a 20 mol% de 3HHx. Os copolímeros assim produzidos foram extraídos e analisados quanto às propriedades físicas. A análise de fluxos metabólicos indicou que a produção de PHA pode ser aumentada com mudanças no metabolismo central e deleção/superexpressão de genes. / The biodegradable copolymer poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) P(3HB-co-3HHx) is a polyhydroxyalkanoate (PHA) presenting various applications. The bacterium Burkholderia sacchari accumulated P(3HB-co-2mol%3HHx) from glucose and hexanoic acid. In order to obtain P(3HB-co-3HHx) with different 3HHx amounts by B. sacchari, recombinant strains containing phaPCJ operon genes from Aeromonas spp were constructed. Recombinant strains produced P(3HB-co-3HHx) from hexanoic, lauric and linoleic acids, with contents of 3HHx ranging from 1.88 to 18 mol%. Experiments with the recombinant in bioreactor, fed in the accumulation phase by glucose 140 g.l-1and hexanoic acid 0-45 g.l-1, resulted in copolymers with compositions ranging from 0 to 20 mol% of 3HHx. The copolymers produced were extracted and analyzed for physical properties. The metabolic flux analysis indicated that PHA production can be increased by modifying the central metabolism and deleting/ overexpressing genes.

O copolímero biodegradável poli(3-hidroxibutirato-co-3-hidroxihexanoato) P(3HB-co-3HHx) é um polihidroxialcanoato (PHA) que apresenta várias aplicações. A bactéria Burkholderia sacchari acumula P(3HB-co-2mol%3HHx), a partir de glicose e ácido hexanoico. Com o objetivo de obter P(3HB-co-3HHx) com diferentes teores de 3HHx por B. sacchari, foram construídas linhagens recombinantes, contendo genes do operon phaPCJ de Aeromonas spp. Os recombinantes produziram P(3HB-co-3HHx), a partir de ácidos hexanoico, láurico e linoleico, com teores de 3HHx entre 1,88-18 mol%. Experimentos em biorreator com o recombinante, alimentada na fase de acúmulo por glicose 140 g/L e ácido hexanoico entre 0-45 g/L, resultaram copolímeros com composições variando de 0 a 20 mol% de 3HHx. Os copolímeros assim produzidos foram extraídos e analisados quanto às propriedades físicas. A análise de fluxos metabólicos indicou que a produção de PHA pode ser aumentada com mudanças no metabolismo central e deleção/superexpressão de genes. / The biodegradable copolymer poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) P(3HB-co-3HHx) is a polyhydroxyalkanoate (PHA) presenting various applications. The bacterium Burkholderia sacchari accumulated P(3HB-co-2mol%3HHx) from glucose and hexanoic acid. In order to obtain P(3HB-co-3HHx) with different 3HHx amounts by B. sacchari, recombinant strains containing phaPCJ operon genes from Aeromonas spp were constructed. Recombinant strains produced P(3HB-co-3HHx) from hexanoic, lauric and linoleic acids, with contents of 3HHx ranging from 1.88 to 18 mol%. Experiments with the recombinant in bioreactor, fed in the accumulation phase by glucose 140 g.l-1and hexanoic acid 0-45 g.l-1, resulted in copolymers with compositions ranging from 0 to 20 mol% of 3HHx. The copolymers produced were extracted and analyzed for physical properties. The metabolic flux analysis indicated that PHA production can be increased by modifying the central metabolism and deleting/ overexpressing genes.

Made available in DSpace on 2016-04-28T20:17:28Z (GMT). No. of bitstreams: 1 2008 - Ronaldo Leao Guimaraes.pdf: 352046 bytes, checksum: 7639df2176f027af3722d0b90e93a856 (MD5) Previous issue date: 2008-12-01 / The aim of this study was to evaluate the presence of Vibrio sp., Aeromonas sp. and Enterobacteriaceae in marine and farmer shrimp. For this, 20 samples of marine shrimp (Penaeus sp.) were collected from Itajuru channel, Cabo Frio, RJ, and other 20 samples of farmer shrimp were acquired in municipal market of Cabo Frio, RJ between January to August 2007. These samples were processed using standard methodology and inoculated in selective media for the research of these microorganisms. The isolates identification was performed by specific biochemical tests. 88 strains were isolated; 51.1% belonging to Vibrio sp., 28.5% to Aeromonas sp. and 20.4% to Plesiomonas shigelloides. It was not detected the presence of E. coli or Salmonella. Among the Vibrio sp., there was a prevalence of Vibrio alginolyticus with 45% and Vibrio parahaemolyticus with 10.5%. Aeromonas hydrophila contributed with 30% of the isolates of this genus and 83.3% of the isolates from Plesiomonas shigelloides were obtained from marine shrimp while 16.7% were obtained from farmer shrimp. The results show a high frequency of potentially pathogenic strains in these shrimps, emphasizing the importance of monitoring these organisms not only to shrimp industry but also to public health. / Este trabalho teve como objetivo pesquisar em camar?es de vida livre e criados em cativeiro a presen?a de microrganismos dos g?neros Vibrio, Aeromonas e representantes da fam?lia Enterobacteriaceae. Para tal, foram coletadas 20 amostras de camar?o de vida livre (Penaeus sp.) do canal de Itajuru, Cabo Frio, RJ; e outras 20 amostras de camar?o de cativeiro do com?rcio local deste munic?pio, no per?odo de Janeiro a Agosto de 2007. Estas amostras foram processadas seguindo metodologia padr?o e inoculadas em meios seletivos para a pesquisa dos microrganismos citados. A identifica??o dos isolados foi realizada atrav?s de testes bioqu?micos espec?ficos. 88 cepas foram isoladas, das quais, 51,1% pertencente ao g?nero Vibrio, 28,5% ao g?nero Aeromonas e 20,4% a Plesiomonas shigelloides. Em nenhuma amostra foi detectada a presen?a de E. coli ou Salmonella spp. Dentre os Vibrios, houve predomin?ncia de Vibrio alginolyticus com 45% e Vibrio parahaemolyticus com 10,5%. Aeromonas hydrophila contribuiu com 30% dos isolados deste g?nero e 83,3% dos isolados de Plesiomonas shigelloides foram obtidos de camar?o de vida livre enquanto 16,7% foram obtidos de camar?o de cativeiro. Os resultados apresentam uma grande freq??ncia de cepas potencialmente patog?nicas ao homem presentes nestes camar?es; ressaltando a import?ncia do monitoramento destes organismos n?o s? para a ind?stria de carcinicultura como tamb?m para a sa?de p?blica.

Aeromonas hydrophila is an opportunistic pathogen that causes a wide range of symptoms and diseases in fish. Development of a commercial vaccine has been problematic due to the heterogenicity between isolates of A. hydrophila. A new approach using immunoproteomics was used in this study to try to develop a vaccine that would protect against a wide range of A. hydrophila strains. The virulence of 14 isolates of A. hydrophila from different geographical regions was determined in common carp (Cyprinus carpio) indicating that 6 isolates were virulent, while 8 isolates were avirulent. Expression of cellular and extracellular products (ECP) of six of these isolates (4 virulent and 2 avirulent isolates) were examined following culture of the bacterium in vitro, in tryptic soy broth, and in vivo, in dialysis tubing placed within the peritoneal cavity of carp. Two types of molecular weight cut off tubes (25 and 100 kDa) were used for the implants. Whole cell (WC), outer membrane protein (OMP) and ECPs of the bacteria grown in vitro and in vivo were analysed by 1 dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis (1D SDS-PAGE). Additionally, 2D SDS-PAGE was used to analyse WC preparations of A. hydrophila grown in vitro and in vivo. The production of unique proteins and up and down-regulation of protein expression were observed in all the preparations of bacteria grown in vitro and in vivo. Unique bands were seen in the 1D SDS-PAGE at 58 and 55 kDa for WC and OMP preparations, respectively, for all the isolates cultured in vivo. Bands of increased intensity were observed at 70, 55, 50 and 25 kDa with WC preparations for the virulent isolates cultured in vivo. Analysis of WC preparations by 2D SDS-PAGE indicated differences in the expression of spots between bacteria cultured in vitro and in vivo. A number of unique spots, mostly between 30 and 80 kDa with pI values ranging from 5.0-6.0 were observed in the bacteria grown in vivo. The protein profiles of different preparations (WC, OMP, ECP) of bacteria cultured in vitro and in vivo were screened by 1D Western blot using antibodies from carp artificially infected with different isolates of A. hydrophila to identify potential vaccine candidates. The WC preparations of A. hydrophila (T4 isolate) grown in vitro were also analysed by 2D Western blot. A 50 kDa protein of A. hydrophila was found to be the most immunogenic molecule in both WC and OMP of bacteria grown both in vitro and in vivo. The protection efficacy of this protein was determined in goldfish by vaccinating fish with electro-eluted 50 kDa protein then challenging the fish with A. hydrophila. Fish were also passively immunised with fish sera raised to the 50 kDa protein and then challenged. The relative percentage survival (RPS) was 67 % in the vaccination trial, while the results were inconclusive for the passive immunisation trial. The 50 kDa protein was confirmed to be the S-layer protein of A. hydrophila following identification using matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS). Recombinant S-layer protein was then produced and the cross-protection efficacy of this protein against six virulent isolates of A. hydrophila was confirmed in a large scale vaccination trial using carp. The RPS value for the 6 isolates of A. hydrophila ranged from between 56 and 87 %. The results of this project suggest that the immunogenic S-layer protein of A. hydrophila could be used as a common antigen to protect fish against infection by different isolates of this pathogenic bacterium.

Diss. (sammanfattning) Uppsala : Sveriges lantbruksuniv., 2002. / Härtill 4 uppsatser.

Thesis (M.S.)--Miami University, Dept. of Chemistry and Biochemistry, 2004. / Title from first page of PDF document. Includes bibliographical references (p. 57-64).

Made available in DSpace on 2014-06-11T19:32:59Z (GMT). No. of bitstreams: 0 Previous issue date: 2013-01-14Bitstream added on 2014-06-13T21:05:30Z : No. of bitstreams: 1 santos_vg_dr_botfmvz.pdf: 418050 bytes, checksum: ff8ea12e28e936c31b32dcbb33f589a4 (MD5) / Avaliou-se a ação imunoestimulante da própolis bruta em dietas para tilápia-do-Nilo submetidas a desafio com a Aeromonas hydrophila. Foram alojados 280 peixes (50 ± 5,7 g) em 35 aquários circulares (250 L), oito peixes/aquário. O delineamento estatístico utilizado foi inteiramente casualizado, caracterizado por sete tratamentos, dieta controle e as demais com suplementação de própolis de 0,5; 1,0; 1,5; 2,0; 2,5 e 3,0% com cinco repetições. Os peixes foram pesados no início e final do experimento para avaliação dos parâmetros de desempenho produtivo. Após trinta dias, 16 peixes de cada tratamento foram anestesiados com benzocaína (1,0g/15L de água) para coleta do sangue e avaliação dos parâmetros hematológicos e imunológicos, caracterizando período anterior ao desafio bacteriano. Os parâmetros hematológicos e imunológicos avaliados foram: número de eritrócitos, porcentagem de hematócrito, taxa de hemoglobina, volume corpuscular médio (VCM), concentração de hemoglobina corpuscular média (CHCM) e peróxido de hidrogênio e óxido nitrico (NO). Em seguida, oito peixes de cada tratamento foram inoculados com injeção intraperitoneal com cultura da bactéria contendo 2,0x107UFC/mL e transferidos para 28 aquários plásticos (40L), dois peixes por aquário. Os mesmos parâmetros foram avaliados após 15 dias. Os resultados demonstraram que níveis elevados de própolis bruta influenciaram na redução do ganho de peso e da hemoglobina e aumento do NO. Comparando-se os momentos, observou-se que após o desafio, os peixes apresentaram valores menores de hematócrito, VCM, neutrófilos e NO e maior CHCM. Concluiu-se que a própolis bruta interferiu no crescimento e na saúde da tilápia-do-Nilo / Immunostimulant effects of crude propolis supplementation on diets to Nile tilapia, submitted to Aeromonas hydrophila, were investigated. Thus, 280 Nile tilapia (initial weight = 50 ± 5,7 g/fish), were allocated in 35 tanks (250 l), eight fish/tank. The experiment was conducted in a completely randomized design, characterized by seven treatments, diet control and the other with propolis supplementation of 0.5, 1.0, 1.5, 2.0, 2.5 and 3.0% with five repetitions. The fish were weighed at the beginning and final of experimental for evaluating the performance parameters. After thirty days, 16 fishes/treatment were anesthetized with benzocaine (1.0g/15L of water), and collected blood to evaluation to haematological and immunological parameters, characterizing the period before to bacterial challenge. The parameters analized were: red blood cell count , haematocrit, haemoglobin, mean corpuscular volume (MCV), mean corpuscular haemoglobin concentration (MCHC), and hydrogen peroxide and nitric oxide (NO). Then, eight fish of each treatment were inoculated with intraperitoneal injection with culture of A. Hydrophila containing 2.0x107UFC/mL, and transferred to twenty eight plastic tanks (40L), and two fishes per tank. The same haematological and immunological parameters were evaluated after fifteen days. The results showed that high levels of crude propolis influence at decrease of weight gain and Hb, an increase of NO. Comparing the moments, was observed that after challenge the fishes showed low value of Htc, MCV, neutrophils and NO, also high value of MCHC. It was concluded that crude propolis affect the growth and health of the-Nile tilapia

Orientador: Margarida Maria Barros / Coorientador: Ricardo de Oliveira Orsi / Banca: Luiz Edivaldo Pezzato / Banca: Lidia Mara Ruv Carelli Barreto / Banca: Maria José Tavares Ranzani de Paiva / Banca: Igo Gomes Guimarães / Resumo: Avaliou-se a ação imunoestimulante da própolis bruta em dietas para tilápia-do-Nilo submetidas a desafio com a Aeromonas hydrophila. Foram alojados 280 peixes (50 ± 5,7 g) em 35 aquários circulares (250 L), oito peixes/aquário. O delineamento estatístico utilizado foi inteiramente casualizado, caracterizado por sete tratamentos, dieta controle e as demais com suplementação de própolis de 0,5; 1,0; 1,5; 2,0; 2,5 e 3,0% com cinco repetições. Os peixes foram pesados no início e final do experimento para avaliação dos parâmetros de desempenho produtivo. Após trinta dias, 16 peixes de cada tratamento foram anestesiados com benzocaína (1,0g/15L de água) para coleta do sangue e avaliação dos parâmetros hematológicos e imunológicos, caracterizando período anterior ao desafio bacteriano. Os parâmetros hematológicos e imunológicos avaliados foram: número de eritrócitos, porcentagem de hematócrito, taxa de hemoglobina, volume corpuscular médio (VCM), concentração de hemoglobina corpuscular média (CHCM) e peróxido de hidrogênio e óxido nitrico (NO). Em seguida, oito peixes de cada tratamento foram inoculados com injeção intraperitoneal com cultura da bactéria contendo 2,0x107UFC/mL e transferidos para 28 aquários plásticos (40L), dois peixes por aquário. Os mesmos parâmetros foram avaliados após 15 dias. Os resultados demonstraram que níveis elevados de própolis bruta influenciaram na redução do ganho de peso e da hemoglobina e aumento do NO. Comparando-se os momentos, observou-se que após o desafio, os peixes apresentaram valores menores de hematócrito, VCM, neutrófilos e NO e maior CHCM. Concluiu-se que a própolis bruta interferiu no crescimento e na saúde da tilápia-do-Nilo / Abstract: Immunostimulant effects of crude propolis supplementation on diets to Nile tilapia, submitted to Aeromonas hydrophila, were investigated. Thus, 280 Nile tilapia (initial weight = 50 ± 5,7 g/fish), were allocated in 35 tanks (250 l), eight fish/tank. The experiment was conducted in a completely randomized design, characterized by seven treatments, diet control and the other with propolis supplementation of 0.5, 1.0, 1.5, 2.0, 2.5 and 3.0% with five repetitions. The fish were weighed at the beginning and final of experimental for evaluating the performance parameters. After thirty days, 16 fishes/treatment were anesthetized with benzocaine (1.0g/15L of water), and collected blood to evaluation to haematological and immunological parameters, characterizing the period before to bacterial challenge. The parameters analized were: red blood cell count , haematocrit, haemoglobin, mean corpuscular volume (MCV), mean corpuscular haemoglobin concentration (MCHC), and hydrogen peroxide and nitric oxide (NO). Then, eight fish of each treatment were inoculated with intraperitoneal injection with culture of A. Hydrophila containing 2.0x107UFC/mL, and transferred to twenty eight plastic tanks (40L), and two fishes per tank. The same haematological and immunological parameters were evaluated after fifteen days. The results showed that high levels of crude propolis influence at decrease of weight gain and Hb, an increase of NO. Comparing the moments, was observed that after challenge the fishes showed low value of Htc, MCV, neutrophils and NO, also high value of MCHC. It was concluded that crude propolis affect the growth and health of the-Nile tilapia / Doutor