Submitted by FERNANDA DA SILVA VON PORSTER (fdsvporster@univates.br) on 2014-09-29T17:42:22Z No. of bitstreams: 3 license_text: 22302 bytes, checksum: 1e0094e9d8adcf16b18effef4ce7ed83 (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) 2014LuisAlbertoPereiraMachado.pdf: 1981082 bytes, checksum: 5d67114fb9c3571e082ce983723758e1 (MD5) / Approved for entry into archive by Ana Paula Lisboa Monteiro (monteiro@univates.br) on 2014-10-06T14:03:17Z (GMT) No. of bitstreams: 3 license_text: 22302 bytes, checksum: 1e0094e9d8adcf16b18effef4ce7ed83 (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) 2014LuisAlbertoPereiraMachado.pdf: 1981082 bytes, checksum: 5d67114fb9c3571e082ce983723758e1 (MD5) / Made available in DSpace on 2014-10-06T14:03:17Z (GMT). No. of bitstreams: 3 license_text: 22302 bytes, checksum: 1e0094e9d8adcf16b18effef4ce7ed83 (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) 2014LuisAlbertoPereiraMachado.pdf: 1981082 bytes, checksum: 5d67114fb9c3571e082ce983723758e1 (MD5) / A carne suína representa importante fonte de proteína animal para o mundo, porém vem sendo associada a surtos de toxinfecção alimentar. Uma das causas destes surtos é a contaminação por Escherichia coli (E. coli), encontrada no trato intestinal e ambiente dos suínos abatidos para produção de carnes in natura e industrializadas. No contexto de segurança alimentar, os surtos por Escherichia coli produtoras de toxina Shiga (STEC) são os melhores documentados. No mundo, diversos surtos causados por ingestão de alimentos de origem animal já foram documentados, porem no Brasil, existem poucos dados sobre a ocorrência de STEC em eventos de toxinfecção alimentar, bem como a presença nos animais e, consequentemente, na carne suína. O objetivo deste trabalho foi quantificar a contaminação por E. coli de carcaças suínas abatidos em frigoríficos comerciais localizados nos estados da Região Sul do Brasil, e identificar por Reação em Cadeia da Polimerase (PCR) a presença de E. coli produtora das toxinas stx1 e eaeA. Foram realizados swabs de 272 carcaças suínas em abatedouros frigoríficos localizados nos estados do RS, SC e PR. As contaminações por E. coli foram identificadas em 25 carcaças, sendo 20 estabelecimento do Rio Grande do Sul, cinco no do Paraná e nenhuma amostra positiva na coleta em Santa Catarina. Das amostras positivas foram extraídos DNA para genotipagem por PCR. Nenhuma amostra apresentou o gene stx1, porém o gene eaeA foi identificado em 13 amostras, nas diferentes regiões da carcaça. A identificação da incidência dos genes eaeA e stx1 nas carcaças pela técnica de PCR pode ser uma ferramenta útil no rastreamento da contaminação bacteriana ao longo dos processos do abatedouro, podendo auxiliar na redução de casos de toxinfecções alimentares causadas por E. coli e outros microrganismos.

Dissertação (mestrado)—Universidade de Brasília, Faculdade de Agronomia e Medicina Veterinária, Programa de Pós-Graduação em Saúde Animal, 2016. / Micro-organismos patogênicos podem contaminar os alimentos por meio da manipulação, higienização e controle ambiental insatisfatórios. Além da adoção das Boas Práticas, o uso de antimicrobianos em alimentos constituiu um passo importante para o controle de doenças infecciosas. Nos sistemas em que ocorre migração do composto ativo para o alimento devem ser considerados apenas aqueles que são aprovados como aditivos alimentares. O objetivo deste estudo foi avaliar a influência da concentração de sal na ação da natamicina sobre micro-organismos patogênicos visto que este conservante tem sido utilizado em banhos de imersão em diversos laticínios no país. Cepas de Candida albicans, Escherichia coli e Staphylococcus aureus foram inoculadas em diferentes concentrações salinas e em água peptonada e receberam tratamentos com natamicina. Estas soluções foram mantidas a temperatura de 12°C e o comportamento dos micro-organismos avaliados com 0, 24 e 48 horas (T0, T1 e T2). Cada micro-organismo foi avaliado isoladamente bem como a associação de C. albicans e E. coli. Nas condições propostas pela pesquisa, foi possível concluir que a natamicina 0,025% não apresenta eficácia sobre Candida albicans inoculada em concentrações salinas abaixo de 5%. Os resultados obtidos nas contagens de E. coli sugerem que a natamicina pode interferir no seu desenvolvimento mesmo em concentrações que podem ser consideradas baixas (0,1%) e em condições de salinidade de 7,5% a 10%. A associação da natamicina com cloreto de sódio potencializa a sua ação antimicrobiana podendo representar economia e o seu uso ser ampliado pelas indústrias. / Pathogenic microorganisms can contaminate food through manipulation cleaning and environmental control unsatisfactory. Besides the adoption of good practice, the use of antimicrobials in food established an important step for the control of infectious diseases. In systems which there are migration from the active ingredient to the food they should be regarded only those compounds that are approved as food additives. The purpose of this study was to evaluate the influence of the concentration of salt in the action of natamycin over pathogenic micro-organisms since this preservative has been used in immersion baths in several dairy products in the country. Strains of Candida albicans, Escherichia coli and Staphylococcus aureus were inoculated in different salt concentrations and in peptone water and were treated with natamycin. These solutions were maintained at temperature of 12 ° C and the behavior of micro-organisms assessed at 0, 24 and 48 hours (T0, T1 and T2). Each microorganism was evaluated singly as well as the combination of C. albicans and E. coli. As proposed by the survey it was concluded that the 0.025 % natamycin has no effect on Candida albicans inoculated in salt concentrations below 5 %. The results obtained from the E. coli counts suggests that natamycin can interfere with their development even at concentrations that may be considered low ( 0.1 %) and saline conditions of 7.5 % to 10 %. The combination of natamycin with sodium chloride enhances its antimicrobial activity may represent the economy and its use be extended by industries.

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Orientador: José Moacir Marin / Banca: Clóvis Wesley Oliveira de Souza / Banca: José Eduardo Zaia / Banca: Alessandra Aparecida Medeiros / Banca: Lúcia Maria Carareto Alves / Resumo: Escherichia coli é a bactéria mais comum entre os patógenos entéricos causadores de doenças intestinais. As diferentes classes de E. coli causadoras de diarréia são reconhecidas através dos fatores de virulência que elas apresentam. As E. coli produtoras de Shiga toxina (STEC), especialmente o sorotipo O157:H7 tem sido associado a diversas doenças no ser humano. Além do sorotipo O157:H7, vários outros sorogrupos não-O157 também estão associados a infecções em humanos. Estas bactérias podem ser recuperadas de muitos animais, mas o gado bovino é reconhecido como o seu mais importante reservatório natural. Para análise da sobrevivência de cepas STEC não-O157 em sistemas de compostagem, inicialmente foram coletadas fezes de três vacas saudáveis que apresentaram E. coli portando o gene stx2, característico de cepas STEC. Foram montados dois sistemas de compostagem: o primeiro foi realizado em vala de 60cmd, no qual E. coli apresentando o gene stx2 foi eliminada após 8, 25 e 30 dias nas temperaturas de 40, 42 e 38°C, respectivamente; o segundo sistema foi realizado sobre o solo em um monte em forma de pirâmide com 1md, no qual as bactérias foram eliminadas após 4, 4 e 7 dias nas temperaturas de 65, 56 e 52°C, respectivamente. A temperatura alcançada durante a compostagem e os microrganismos presentes no esterco parecem ser os responsáveis pela eliminação do patógeno nos sistemas de compostagem, o qual pode ser útil para a redução da carga patogênica presente no esterco destinado para aplicações no solo. / Abstract: Escherichia coli is the most common bacteria among the enteric pathogens able to cause intestinal disease. Several classes of diarrhea-causing E. coli are recognized on the basis of their virulence factors production. Shiga-like toxigenic E. coli (STEC), especially serotype O157:H7, have been associated with many diseases in human beings. Besides sorotype O157:H7, many others non-O157 sorogroups have also been associated with human infections. These bacterias can be isolated from a range of animals, but cattle is generally recognized as the major natural source. To analyze the survival of non-O157 STEC strains in composting system, first was collected faeces from three healthy cows that contain E. coli STEC cells carrying the stx2 gene. Two composting systems were used: the first one was a cave with 60cmd were the E. coli STEC cells with stx2 gene were eliminated after 8, 25 and 30 days at 40, 42 and 38°C, respectively; the second one was a heap pyramid system with 1md, where the cells were eliminated after 4, 4, 7 days at 65, 56 and 52°C, respectively. The reached temperature in the composting systems and the indigenous microorganisms present in the manure seems to contribute to pathogen elimination, what may be a useful means of reducing the pathogen load of manure destined for soil application. / Doutor

Thesis (Ph.D.)--Boston University / PLEASE NOTE: Boston University Libraries did not receive an Authorization To Manage form for this thesis or dissertation. It is therefore not openly accessible, though it may be available by request. If you are the author or principal advisor of this work and would like to request open access for it, please contact us at open-help@bu.edu. Thank you. / Cross reactivation (CR) is defined as the rescue of genetic markers from inactive bacteriophage particles by viable bacteriophage particles. UV was used as the inactivating agent in the experiments reported in this dissertation. Escherichia coli BB was used as the host bacterium and the bacteriophage used was T4. E. coli BB was infected half with UV irradiated T4 and half with normal, unirradiated T4. Under the conditions of the experiment lysis of the infected culture did not take place until 75 minutes after infection. At specific times after infection, infected cells were broken open and the number of intracellular phage per infected cell was determined. The results indicate that a normal size replicating pool of phage molecules is reached as quickly under the conditions of this experiment as when cells are infected with live phage particles. Moreover, this experiment provides further evidence that UV lesions do not replicate. E. coli BB was infected with two different T4 rII mutants. One of these mutants had been UV irradiated (50 lethal hits per phage) and its concentration was adjusted so that no cell received more than one of these particles. The other mutant was unirradiated and each cell received 2 to 3 of these particles. The dose of irradiation chosen was such that essentially all the irradiated particles had at least one lesion between the two loci. Until 22 minutes after infection, infected cells were plated on streptomycin medium to break them open and on a normal medium to allow natural lysis to occur. A comparison of counts on the two types of media showed the fraction of cells undergoing CR which had done so by that time. The results of this experiment show that CR is completed at the end of one normal latent period, i.e., before normal lysis. This is so despite the fact that at the dose of UV employed, only 4.8 per cent of the cells infected with both mutants showed CR. The cells which did show CR did not show an increase in the frequency of CR recombinants upon extension of the latent period. An increase in this frequency would have been expected if many copies of unirradiated portions of irradiated phage genomes had been present in the phage replicating pool. The results of these last two sets of experiments are interpretable as a failure of unhit portions of UV irradiated phage genomes to replicate normally within the phage replicating pool. Somehow these pieces of genome are removed from the pool faster than they are synthesized. It is noteworthy that these results are also compatible with the hypothesis that at the dose of UV employed, unirradiated portions of irradiated phage genomes do not replicate. A model of CR is presented which is consistent with present ideas of bacteriophage T4 genetic recombination, as well as with CR data from other sources. Moreover, this model requires no replication of unirradiated portions of irradiated phage DNA genomes. Finally, the phenomenon of lysis inhibition, as manifested by extension of the latent period by super infection of infected cells under appropriate conditions has been examined. It was found that more and more of these infected cells which had been almost completely deprived of a constant source of superinfection lysed after having been lysis inhibited only once. These results show that continuous reinfection is required to maintain lysis inhibition. / 2031-01-01

Thesis (Ph.D.)--Massachusetts Institute of Technology, Dept. of Biology, 1981. / MICROFICHE COPY AVAILABLE IN ARCHIVES AND SCIENCE. / Bibliography: leaves 174-182. / by Daniel Jeffrey Capon. / Ph.D.

Le réseau de protéines chaperons est hautement conservé dans l'ensemble du vivant. Il régule l'homéostasie des protéines au sein de la cellule en condition de croissance normale ainsi qu'en réponse à des stress environnementaux. Les chaperons membres de la famille HSP70 (Heat Shock Protein 70 kDa), famille particulièrement conservée, agissent tout au long de la biogénèse des protéines et orchestrent une pléthore de processus cellulaires liés au repliement et/ou au remodelage de protéines. Le cycle ATP-dépendant du chaperon HSP70 repose sur une étroite collaboration avec ses partenaires co-chaperons. Parmi ces co-chaperons, on distingue les membres de la famille DnaJ/HSP40 qui transfèrent les substrats vers HSP70 et stimulent son activité ATPasique, et les facteurs d'échange de nucléotides qui assurent la réinitialisation du cycle d'HSP70 permettant ainsi la libération du substrat. Au sein de la bactérie E. coli, la protéine HSP70 est appelée DnaK. Elle agit de concert avec les deux co-chaperons DnaJ et GrpE (ensemble nommés DnaKJE) afin d'assister les protéines dans leur repliement au cours de la synthèse de novo, de désagréger des protéines mal repliées, de faciliter l'adressage et le passage de protéines à travers les membranes biologiques, et de remodeler certains complexes protéiques impliqués dans des processus cellulaires variés. DnaKJE coopère efficacement avec d'autres systèmes chaperons majeurs, tels que la protéine Trigger Factor (TF) associée aux ribosomes et le complexe chaperonine GroESL, notamment pour le repliement de protéines nouvellement synthétisées dans le cytosol. De plus, une des fonctions cellulaires majeure du système DnaKJE est son implication dans la réponse au stress thermique (Heat Shock Response - HSR). DnaKJE contrôle la HSR en interagissant directement avec le facteur de transcription s32, sous-unité de l'ARN polymérase. Cette interaction facilite la dégradation de s32 par la protéase FtsH. En condition de stress, l'accumulation de protéines mal repliées au sein de la cellule entraine le recrutement de DnaKJE et par conséquent, la stabilisation de s32. Suite à cette stabilisation, une induction de la transcription de plus d'une centaine de gènes codant entre autres, pour des protéines chaperons et des protéases se met en place dans la cellule pour lutter contre le stress environnant. De ce fait, DnaK et ses co-chaperons sont considérés comme des éléments clés de la réponse cellulaire contre le collapse de l'homéostasie protéique par action directe sur des protéines mal repliées et indirecte en modulant la synthèse de nombreuses HSPs, via s32. L'étude récente de l'intéractome de DnaK révèle qu'au moins 50% des enzymes impliquées au sein du métabolisme central (MC) de la cellule interagissent avec DnaK à température physiologique. A travers l'analyse d'une banque de suppresseurs multi-copie, nous avons identifié six gènes associés au MC : ackA, ldhA, lpd, pykF, talB et csrC qui lorsqu'ils sont surexprimés, permettent de restaurer partiellement le défaut de croissance d'une souche mutante n'exprimant pas les chaperons DnaK et Trigger Factor (deltatig deltadnaKJ). Remarquablement, la surexpression d'ackA, talB et csrC supprime également le défaut de croissance d'un mutant dnaK à haute température, ce qui suggère une implication importante de DnaK au niveau du MC. Dans ce projet, l'implication de DnaK dans le fonctionnement du métabolisme carboné a été établi par une analyse métabolique combinant analyses macro-cinétiques (suivi de croissance, analyse de la consommation des substrats et de la production de produits du métabolisme) sur différentes sources de carbones seules ou en mélange et analyses micro-cinétiques (flux métaboliques par marquage 13C). Finalement, ces travaux apportent différentes hypothèses quant au rôle de DnaK dans le contrôle du MC, directement ou indirectement via la régulation de la HSR, en réponse à une défaillance de l'homéostasie protéique ou d'une carence nutritionnelle. / Intricate networks of highly conserved molecular chaperone machines govern cellular protein homeostasis, both under lenient and more stressful growth conditions. Members of the highly conserved HSP70 family of molecular chaperones are key players in this process, acting at nearly every step in protein biogenesis. The ATP-dependent chaperone cycle of HSP70 chaperones relies upon the cooperation with a cohort of essential cochaperones, including DnaJ/HSP40 family members that recruit the chaperone to specific substrate and/or cellular localization and stimulate its ATPase activity, and nucleotide exchange factors, which insure proper resetting of the chaperone cycle and the resulting substrate release. In the bacterium Escherichia coli, the multifunctional HSP70 chaperone, named DnaK, acts in concert with its cochaperones DnaJ and GrpE (all together referred as DnaKJE) to efficiently, assist de novo protein folding, protein disaggregation, protein targeting and translocation through biological membranes, and protein complexes remodeling leading to multiple cellular activities. Remarkably, previous works also showed that DnaKJE can efficiently cooperate with other major cytosolic chaperones, including the ribosome-bound Trigger Factor (TF) and the chaperonin GroESL, especially during the folding of newly-synthesized cytosolic proteins. In addition, one of the key cellular functions of DnaKJE in E. coli is the regulation of the heat shock response (HSR). In this case, DnaKJE controls the HSR by interacting directly with the heat shock sigma factor s32 subunit of the RNA polymerase to facilitate it degradation by the FtsH protease. Under stress condition, DnaKJE is recruited to accumulating misfolded proteins, leading to an increased stability of s32 and the subsequent induction of more than hundred heat shock proteins. Therefore, DnaK, and its cochaperones are central components of the cellular response to proteostasis collapse, both by acting directly on misfolded proteins and by modulating the synthesis a plethora of heat shock chaperones and proteases. The recently described in vivo interactome of DnaK in E. coli revealed that at least 50% of the central metabolism enzymes interact with DnaK at physiological temperature. Remarkably, through a multicopy suppression analysis we have now identified six genes of the central metabolism (CM), namely ackA, ldhA, lpd, pykF, talB and csrC, which when overexpressed partially suppress the growth defect of the sensitive double mutant lacking DnaK and Trigger Factor (deltatig deltadnaKJ ), with half of them, namely ackA, talB and csrC, additionally suppressing the growth defect of the single ?dnaKJ mutation at high temperature, thus strongly suggesting a major role of DnaK in this process. Using a combination of growth assays on specific carbon sources entering the CM at various metabolic nodes with NMR analyses for characterizing the carbon source assimilation, identifying and quantifying the metabolism by-products and determining metabolic flux rearrangements, we show that DnaKJE impacts the responsiveness of the central metabolism by acting either directly at the level of the CM or along the first step of substrate assimilation. How does the multifunctional DnaK chaperone modulate the CM, either directly or indirectly via the control of the HSR, in response to proteostasis failure or nutrient starvation is discussed.

O consumo de vegetais e de frutas tem aumentado mundialmente, bem como os surtos alimentares envolvendo esses alimentos, especialmente a alface que é o vegetal folhoso mais consumido em nível mundial. Dessa forma, o objetivo desse estudo foi realizar uma avaliação quantitativa do risco de infecções causadas por Salmonella spp. e por Escherichia coli O157:H7 a partir do consumo de alface produzida e consumida no Rio Grande do Sul, visto que esses patógenos são os mais relacionados a surtos alimentares envolvendo vegetais folhosos em nível mundial. Para melhor compreender o comportamento desses patógenos na alface, eles foram inoculados nesse vegetal separadamente e armazenados sob condições isotérmicas de 5 a 40°C para Salmonella e de 5 a 42ºC para E. coli O157:H7, bem como sob condições não isotérmicas, simulando temperaturas encontradas da colheita até a venda da alface no Rio Grande do Sul. Dados experimentais demonstraram que ambas as bactérias podem se multiplicar em todas as temperaturas examinadas. Também foi proposto um parâmetro de tempo de multiplicação insignificante (ς), o qual fornece o tempo em que a alface pode ser exposta a uma temperatura específica e não apresentar uma multiplicação expressiva. O ς foi desenvolvido com base na equação do modelo primário de Baranyi e no conceito do potencial de crescimento. ς é o valor da fase lag adicionado do tempo necessário para população microbiana aumentar 0,5 log UFC/g. O ς da alface exposta a 37 °C foi de 1,3 h, enquanto que a 5 °C foi de 3,3 dias. Além dos modelos adequados, dados de prevalência e concentração são primordiais na avaliação de risco. Assim, foi realizada uma revisão sistemática da literatura para buscar esses dados A prevalência mundial encontrada foi de 0,041 para ambos os patógenos na alface. Já a prevalência dos países desenvolvidos foi de 0,028 para Salmonella e de 0,125 para E. coli (EHEC), enquanto que nos países em desenvolvimento foi de 0,064 para Salmonella e 0,024 para E. coli (EHEC). A concentração de Salmonella em alface, em países em desenvolvimento, variou de 4,57 a 218,78 NMP/g, e para E. coli (EHEC) a concentração foi de < 3,0 NMP/g até > 1100 NMP/g. O modelo de avaliação quantitativa de risco microbiológico foi composto por nove módulos, desde o armazenamento da alface nas fazendas produtoras até o consumo. O risco médio (baseado no cenário mais comumente encontrado no Rio Grande do Sul) de infecção por Salmonella por mês foi de 0,017, enquanto que por E. coli O157:H7 foi de 0,006. Assim, de modo geral, o risco de infecção por Salmonella é maior do que por E. coli O157:H7 quando a alface é produzida e consumida nesse estado. Todos os cenários alternativos à correta higienização da alface (lavar as folhas de alface com água potável seguido de imersão em 200 ppm de cloro livre, por 15 minutos e enxaguar com água potável) aumentaram o risco. A principal redução do risco foi identificada no cenário que considerou o uso de refrigeração em todos os módulos do modelo. Análises de sensibilidade indicaram que, além da manutenção da cadeia fria e do procedimento correto de higienização, é importante reduzir a prevalência e a concentração dos patógenos na alface, a fim de diminuir o risco de infecção por essas bactérias. Por fim, a avaliação de risco desenvolvida nessa tese pode auxiliar no desenvolvimento de estratégias de intervenção para mitigar esse risco. / The consumption of vegetables and fruits has increased worldwide, as well as foodborne outbreaks involving these foods, especially lettuce that is the most consumed leafy vegetable in the world. Thus, the objective of this study was to carry out a quantitative microbial risk assessment of Salmonella spp. and Escherichia coli O157: H7 on lettuce produced and consumed in Rio Grande do Sul, since these pathogens are the most related to foodborne outbreaks involving leafy vegetables worldwide. To study the behavior of these pathogens on lettuce, they were inoculated on this vegetable separately and stored under isothermal conditions of 5 to 40 °C for Salmonella and 5 to 42 °C for E. coli O157:H7, as well as under non-isothermal conditions, simulating temperatures from the harvest until the sale of lettuce in Rio Grande do Sul. Experimental data demonstrated that both bacteria can growth at all temperatures examined. A negligible growth time parameter (ς) has also been proposed, which provides the time that lettuce can be exposed to a specific temperature and does not present an expressive growth. The ς was developed based on the equation of the Baranyi primary model and the concept of growth potential. ς is the lag phase added value of the time required for microbial population to increase 0.5 log CFU/g. The ς of lettuce exposed at 37 ºC was 1.3 h, whereas at 5 ºC it was 3.3 days. In addition, prevalence and concentration data are paramount in the risk assessment studies. Thus, a systematic review of the literature was carried out to collect these data. The global prevalence found was 0.041 for both pathogens in lettuce The prevalence of developed countries was 0.028 for Salmonella and 0.125 for E. coli (EHEC), while in developing countries it was 0.064 for Salmonella and 0.024 for E. coli (EHEC). The concentration of Salmonella in lettuce in developing countries ranged from 4.57 to 218.78 MPN/g, and for E. coli (EHEC) the concentration was < 3.0 MPN/g to > 1100 MPN/g. The quantitative microbial risk assessment model was composed by nine modules, from lettuce storage on farms to consumption. The average risk (based on the scenario most commonly found in Rio Grande do Sul) of Salmonella infection per month was 0.017, whereas for E. coli O157:H7 it was 0.006. Thus, in general, the risk of infection by Salmonella is higher than by E. coli O157:H7 when lettuce is produced and consumed in this State. All scenarios that were alternative to the correct hygiene of lettuce (washing lettuce leaves with drinking water followed by immersion in 200 ppm of free chlorine for 15 minutes and rinsing with potable water) increased the risk. The main risk reduction was identified in the scenario that considered the use of refrigeration in all modules of the model. Sensitivity analyzes indicated that, in addition to maintaining the cold chain and the correct hygienization procedure, it is important to reduce the prevalence and concentration of pathogens in lettuce, in order to reduce the risk of infection by these bacteria. Finally, the risk assessment developed in this thesis can help in the development of intervention strategies to mitigate this risk.

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Publicación a texto completo no autorizada por el autor / Determina la correlación entre la presencia de genes de virulencia específicos y su expresión fenotípica en Escherichia coli diarreogénicas, mediante PCR múltiple usando una combinación de 6 pares de cebadores específicos para E.coli diarreogénicas y un medio selectivo nuevo diferencial para el aislamiento de E-coli STEC noO157, EPEC y ETEC. De 100 cepas analizadas, el 31 % correspondieron a E.coli EPEC atípicas, mientras el 5 % fueron típicas porque evidenciaron la presencia del gen eae, y genes eae + bfp respectivamente. De las cepas STEC el 32 % presentaron el gen Stx1, 4 % el gen Stx1 +eae y solo 2 % el gen Stx1+Stx2 y eae. En ETEC el 24 % poseían el gen est que codifica para la enterotoxina termoestable (ST) y 1% poseía los genes est y elt. Los ensayos para determinar la expresión fenotípica de Escherichia coli con genes de virulencia en medios de cultivo diferenciales de Posse nos permitieron identificar Escherichia coli STEC O26 (15%) que crecieron dando colonias purpura como consecuencia de fermentar sacarosa y sorbosa, O111 (14 %) y O103 (1 %) dieron colonias azul purpura por la fermentación de sacarosa y O114 (9 %) que no fermentan sorbosa, así como los serotipos EPEC (36 %) y ETEC (25 %) tampoco fermentaron sorbosa dando colonias rojas. Se determinó que no existe correlación entre la cepas STEC sorbosa negativas y la presencia de genes para la producción de toxinas Shx, así como la presencia de genes est y elt para la producción de enterotoxinas ST y LT en cepas ETEC demostrando que el método de la sorbosa puede ser un marcador fenotípico para la rutina en los laboratorios para la selección de estos patógenos en productos alimenticios, muestras clínicas humanas y de animales. / Tesis