Submitted by Marcelo Andrade Silva (marcelo.andradesilva@ufpe.br) on 2015-03-09T14:23:51Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) DISSERTAÇÃO Catarina Fernandes de Freitas.pdf: 2589861 bytes, checksum: 3f587b558e8abc1410528aa38d8eeb1a (MD5) / Made available in DSpace on 2015-03-09T14:23:51Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) DISSERTAÇÃO Catarina Fernandes de Freitas.pdf: 2589861 bytes, checksum: 3f587b558e8abc1410528aa38d8eeb1a (MD5) Previous issue date: 2014 / A crescente incidência de isolados de Klebsiella pneumoniae resistentes a carbapenêmicos e outras classes de antibióticos -lactâmicos, em casos de infecções relacionadas a assistência a saúde, tem sido amplamente relacionada com a produção de β-lactamases. A necessidade da utilização de dispositivos invasivos como cateteres, em pacientes hospitalizados pode favorecer a colonização destes por diferentes espécies bacterianas, propiciando a formação de biofilmes, o que pode agravar o estado clínico do paciente prejudicando a eficácia terapêutica. O objetivo deste estudo foi detectar a presença dos genes de resistência blaSHV, blaTEM, blaCTX-M, blaKPC; genes de virulência para produção de cápsula polissacarídica (cps), fímbrias do tipo 1 (fimH) e tipo 3 (mrkD); e a capacidade de formação de biofilme por isolados de K. pneumoniae multidroga-resistente (MDR) na ausência e presença de antibióticos. Foram analisados nove isolados de K. pneumoniae oriundos de pacientes hospitalizados na cidade de Recife-PE. A concentração inibitória mínima dos antibióticos cefotaxima, ceftazidima e amicacina foi determinada pelo método de macrodiluição. Foi feita a tipagem molecular através da técnica de amplificação de sequências intergênicas repetitivas de enterobactérias (ERIC-PCR). A determinação da presença dos genes foi realizada por PCR convencional. A formação de biofilme foi analisada após inoculação e incubação dos isolados em meio de cultura com um fragmento de catéter na presença e neumoni dos antibióticos cefotaxima, ceftazidima e amicacina, além da combinação da ceftazidima e amicacina para os que foram resistentes à amicacina. Os neumonia foram processados para microscopia eletrônica de varredura. A genotipagem por ERIC-PCR demonstrou perfis genéticos distintos para os nove isolados. Sete dos nove isolados foram positivos para os genes blaTEM, blaSHV e blaKPC e seis para o gene blaCTX-M. Todos os isolados foram positivos para o gene cps e mrkD e seis para o gene fimH. Todos os isolados formaram biofilme na ausência de antibióticos. Três isolados não formaram biofilme quando submetidos aos antibióticos testados. Em dois isolados a capacidade de formar biofilme melhorou quando submetidos aos antibióticos. Enquanto que sete dos nove isolados submetidos aos antibióticos, a formação de biofilme foi menor quando comparada com o controle. Embora os isolados sejam resistentes aos antibióticos foi possível observar diminuição do número de células aderidas e a diminuição da formação de biofilme por isolados de K. pneumoniae MDR portadores de diferentes β-lactamases e de fatores de virulência, os quais apresentaram comportamentos distintos em relação a formação de biofilme, dependendo do isolado e do antibiótico, podendo este inibir ou induzir uma melhor formação de biofilme.

Submitted by Ramon Santana (ramon.souza@ufpe.br) on 2015-03-10T14:37:00Z No. of bitstreams: 2 Dissertaçao Rita de Cassia Melo.pdf: 1049570 bytes, checksum: 2912f081df833939c5b2ca15a160932c (MD5) license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) / Made available in DSpace on 2015-03-10T14:37:01Z (GMT). No. of bitstreams: 2 Dissertaçao Rita de Cassia Melo.pdf: 1049570 bytes, checksum: 2912f081df833939c5b2ca15a160932c (MD5) license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Previous issue date: 2013-03-15 / Klebsiella pneumoniae é um patógeno oportunista frequentemente associado a infecções hospitalares do trato respiratório e do trato urinário de indivíduos imunocomprometidos e neonatos, e podem produzir diferentes tipos de fatores de virulência, como adesinas fimbriais (genes fimH e mrkD ) e sideróforos, como a yersiniabactina (gene irp2), importantes no desenvolvimento da infecção. O objetivo do presente estudo foi determinar a ocorrência dos genes de virulência fimH, mrkD e irp2 em isolados de K. pneumoniae produtores e não produtores de KPC, provenientes de pacientes de diferentes hospitais de Recife-PE, como também a relação clonal, através da ERIC-PCR, e o perfil de susceptibilidade a antimicrobianos desses isolados bacterianos. Para esse estudo foram selecionados 23 isolados produtores de KPC e 23 isolados não produtores de KPC, todos da espécie K. pneumoniae. Os genes de virulência foram detectados através da PCR e sequenciamento de DNA. Analisando o perfil de susceptibilidade aos antimicrobianos dos isolados selecionados, observamos que a amicacina (n=39/ 84,78%) e a polimixina (n=41/ 89,13%), foram os antimicrobianos de melhor atividade para inibir a K. pneumoniae, tanto KPC-positivas quanto negativas, onde observamos que 5 isolados apresentaram resistência a polimixina, sendo 3 no grupo KPC-positivo e 2 no grupo KPC-negativo. Esse é o primeiro relato da resistência de K. pneumoniae à polimixina. No grupo KPC-positivo foi observada uma alta taxa de resistência à cefalosporinas, seguidas dos carbapenêmicos. Ficou evidenciado que o ertapenem é o melhor antimicrobiano para detectar resistência fenotípica ao grupo dos carbapenêmicos. A tipagem pela ERIC-PCR gerou 37 perfis genéticos, demonstrando que houve Uma disseminação multiclonal de isolados de K. pneumoniae em Recife-PE, Brasil. Dentre os 46 isolados analisados pela ERIC-PCR, cincoperfis agruparam mais de um isolado bacteriano com relação clonal. No presente estudo não foi possível detectar a relação direta entre a presença do gene blaKPC com cada gene de virulência individualmente, visto que os grupos estudados, KPC-positivo e KPC-negativo, apresentaram presenças semelhantes dos genes de virulência. Por outro lado, foi observado que os genes de virulência irp2, mrkD e fimH apresentaram-se juntos com uma maior frequência no grupo KPC-positivo. O acúmulo de genes de virulência em isolados de K. pneumoniae KPC positivos, observado nesse estudo, juntamente com a multirresistência, impõe relevantes limitações terapêuticas no tratamento de infecções causadas por K. pneumoniae em Recife-PE, Brasil.

The work described in this thesis forms part of an international programme concerned with the structure elucidation of the capsular antigens of some Enterobacteriaceae. Many of the Klebsiella and some of the Escherichia coli are pathogenic to man and, hence, they are of interest. The virulence of bacteria is a multifactorial phenomenon, in which characteristic traits of bacteria and their hosts play comparable and complementary roles. It is accepted that pathogens are more virulent when encapsulated, because, nearly all disease causing bacteria have a capsule when freshly isolated from the host. This increase in pathogenicity is related, in part, to the capsular polysaccharides' ability to avoid or attenuate the host defence mechanisms. In the majority of cases the protective aspects of the capsule are overcome in the latter stages of infection when the formation of specific antibodies by the host has occurred. However there are situations in which an immune state of the infected host is virtually never reached, and susceptiblity to the infecting bacteria is maintained even in the advanced stage of an infection. Explanation of this phenomenon becomes possible by analysing the structure of the polysaccharides. The inability of the host to raise an immune response to the capsule may be because the structure of the polysaccharide is similar or identical to the host's carbohydrates. The serological and pathogenic relatedness of encapsulated E. coli and Klebsiella, to the encapsulated strains of other genera, is based on structural identity or similarity of the respective capsules. Capsular polysaccharides are analysed by both chemical and instrumental methods, and, at present, nuclear magnetic resonance spectroscopy is the most important analytical technique

From Introduction: Bacterial cells all have a cytoplasmic membrane (see Figure 1) which regulates the movement of ions and molecules into and out of the bacterium. Enclosing this membrane is a cell wall of which there are two general types, which are differentiated by the Gram stain(02) as being either gram positive or gram negative (depending upon whether they hold the gram stain after washing with ethanol). The cell wall provides the cell with shape and rigidity and is composed, in the case of gram positive types, of peptidoglycan, and in the case of gram negative bacteria, of a peptidoglycan and an outer membrane (see Figure 2). The peptidoglycan layer, common to both cell wall types, consists of a backbone of alternating units of N-acetylglucosamine and N-acetylmuramic acid to which peptides are attached by amide links. This heteropolymer is a highly cross linked mosaic and this gives it strength and rigidity. In gram positive bacteria, this layer also contains two carbohydr ate antigens, a simple polysaccharide and a teichoic acid; these are usually the type specific or major group antigens of the bacterium. Many of the bacteria also produce exopolysaccharides (see Figure 3) either as discrete capsules (for example, the Enterobacteriaceae K antigens) or unattached slime layers (for example, the Enterobacteriaceae M antigens). The vast majority of these polysaccharides are heteroglycans(03) composed of contiguous oligosaccharide repeating units. Their monosaccharide components are largely neutral hexoses, 6-deoxy hexoses and also amino sugars. (03) Pentose units are rare. (03) The capsular polysaccharides usually have a high content of acidic constituents such as uronic acids, phosphate groups, or pyruvate ketals. (01) / KMBT_363 / Adobe Acrobat 9.53 Paper Capture Plug-in

This thesis presents a detailed chemical and spectroscopic determination of the capsular, polysaccharide K-antigen isolated from the Klebsiella bacterium, serotype K43 (culture #2482). The repeating unit of the capsular polysaccharide was found to be of the "3 + 2" repeating unit type. A uronic acid was found as part of a disaccharide side chain and the main chain of the polysaccharide was found to be composed of a neutral trisaccharide of mannose and galactose. The work forms part of an ongoing research interest in bacterial polysaccharides of this laboratory and now completes the structural elucidation of all the Klebsiella K-antigens, bar three antigens which were originally assigned to other laboratories. These data together with the respective serological characteristics of each serotype are available to the molecular biologist, and may result in the production of: vaccine(s) against Klebsiella infections, diagnostic products and novel carrier molecules enabling targeted drug delivery.

A review of the structural studies of bacterial capsular polysaccharides (K-antigens) from Escherichia coli and Klebsiella, and of the trends in modern chemical and instrumental techniques available for the analysis of carbohydrate material is presented. The structural elucidations of the capsular polysaccharides from E. coli K37 and K55, and Klebsiella K39 are reported with comments on the novelty and possible immunological significance of the structures. The usefulness of the bacteriophage degradation technique has been emphasized using the polysaccharides from E. coli K55, and Klebsiella K30 and K39 to demonstrate the scope of the reaction

Seminario de Título entregado a la Universidad de Chile en cumplimiento parcial de los requisitos para optar al Título de Ingeniero en Biotecnología Molecular. / En las últimas dos décadas se ha registrado un incremento significativo en la detección de cepas hipervirulentas y multi-resistentes del patógeno bacteriano Klebsiella pneumoniae, lo cual estaría ligado con el alto dinamismo y la rápida evolución de su genoma. Dentro de los elementos genéticos móviles (EGMs) que determinan dicho dinamismo se encuentran las islas genómicas (IGs), que corresponden a fragmentos de DNA de hasta 200 kb que se integran preferentemente en genes que codifican tRNAs (tDNAs). Se ha demostrado que las IGs de K. pneumoniae y otras enterobacterias portan una gran variedad de genes relacionados con virulencia y resistencia a antibióticos, por lo que su identificación y caracterización a partir del creciente número de genomas secuenciados de esta especie resulta altamente relevante. En esta dirección, el presente trabajo describe el desarrollo de una herramienta para identificar IGs asociadas a tDNAs que se basa en el análisis del contexto genómico de dichos genes, es decir, la organización e identidad de los genes que se encuentran adyacentes en el cromosoma. En una primera etapa, el programa identifica los tDNAs y su contexto para luego clasificarlos y nombrarlos. En una segunda etapa, el programa analiza la continuidad del contexto genómico conservado de cada tDNA y determina la presencia de IGs cuando una disrupción es identificada. El programa creado, se utilizó para analizar el genoma de 50 cepas de K. pneumoniae, donde los resultados obtenidos fueron comparados con la identificación de IGs realizada manualmente y curada en un estudio previo realizado por nuestro grupo. Como resultado, se logró identificar correctamente un 99,3% de todos los tDNAs presentes en dichas cepas, además de 303 IGs de un total de 308 IGs detectadas manualmente. Adicionalmente, el programa mostró un desempeño satisfactorio, aunque mejorable en la detección de integrasas y repetidos directos, detectando ambos elementos en 148 de las IGs identificadas. Finalmente, se comparó el desempeño del programa desarrollado y otros programas para la identificación de IGs en la detección y delimitación de la isla GIE492 caracterizada experimentalmente en un trabajo previo de nuestro grupo, determinando que la detección hecha por el programa desarrollado por nuestro grupo es más sensible y precisa que cualquiera de los otros programas utilizados. / In the last two decades, there has been a significant increase in the detection of hypervirulent and multi-resistant strains of the bacterial pathogen Klebsiella pneumoniae, which would be related to the high dynamism and rapid evolution of its genome. Within the mobile genetic elements (EGMs) that determine this dynamism are Genomic Islands (IGs), which correspond to DNA fragments up to 200 kb that are preferentially integrated into genes encoding tRNAs (tDNAs). It has been shown that IGs from K. pneumoniae and other species from Enterobacteriaceae carry a wide variety of genes related to virulence and antimicrobial drug resistance. Thus, their identification and characterization among the increasing number of sequenced genomes from this species is highly relevant. In this direction, this work describes the development of a bioinformatics tool intended to identify IGs associated to tDNAs based on the analysis of the genomic context of these genes, that is, the organization and identity of the genes that are adjacent to tDNAs in the chromosome. In the first step, the program identifies all the tDNAs and their contexts to classify and name them. In a second step, the program analyzes the continuity of the conserved genomic context for each tDNA, determining the presence of a putative IG when a disruption of such continuity is detected. Our program was used to analyze the genome of a set of 50 K. pneumoniae strains, where the results obtained were compared with the identification of IGs performed and cured manually in a previous study conducted by our group. As a result, 99.3% of all the tDNAs present in the strains were correctly identified. Also, the software predicted 303 IGs from a total of 308 IGs detected manually. Additionally, the program showed a good performance in the identification of integrases and direct repeats, detecting both elements in 148 of the IGs, although there is still room for improvement. Finally, we compared the performance of our software and other programs for the identification of IGs, in the detection and precise delimitation of the xi GIE492 island, which was characterized experimentally in a previous work of our group. We determined that the detection made by our program was more sensitive and precise than any of the other programs tested.

Klebsiella spp. are coliform bacteria that cause mastitis in dairy cattle and account for high mortality rates in infected cows leading to a significant financial loss. Recent outbreaks indicate that within farms a single strain can be responsible for clinical signs in multiple animals. Identification of the virulence of factors enabling Klebsiella spp. survival in the mammary glands of multiple animals may provide insight into host adaptation. In this study, Klebsiella spp. strains were evaluated for their ability to evade neutrophil killing, the primary immune defense in the bovine mammary gland. Our research focused on capsule and biofilm production by Klebsiella spp. when strains were grown in Luria Broth or skim milk to examine the effects on evasion of neutrophil killing. Biofilm production was not significantly related to the ability to resist neutrophil killing nor was capsule (P = 0.29). Farm (P < 0.001), media type (P < 0.005), and strain type by cow (P < 0.001) were found to play significant roles in neutrophil evasion. This suggests farm of origin, media type used, and cow all may play a role in evasion of neutrophils by Klebsiella spp. Further evaluation of virulence factor expression in different media types and the role of individual cow immune responses may provide insight into ability of Klebsiella spp. to cause outbreaks of mastitis in multiple animals. / Master of Science

Submitted by Fabio Sobreira Campos da Costa (fabio.sobreira@ufpe.br) on 2016-08-26T13:01:30Z No. of bitstreams: 3 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) ElzaDissertação: 1942336 bytes, checksum: a9041b50df03d17a3c0538c97db03452 (MD5) ElzaDissertação: 1942336 bytes, checksum: a9041b50df03d17a3c0538c97db03452 (MD5) / Made available in DSpace on 2016-08-26T13:01:30Z (GMT). No. of bitstreams: 3 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) ElzaDissertação: 1942336 bytes, checksum: a9041b50df03d17a3c0538c97db03452 (MD5) ElzaDissertação: 1942336 bytes, checksum: a9041b50df03d17a3c0538c97db03452 (MD5) Previous issue date: 2016-02-24 / CAPEs / Klebsiella pneumoniae e Enterobacter aerogenes têm se destacado como importantes agentes de infecções relacionadas à assistência à saúde (IRAS), causando principalmente infecções de feridas, dos tratos urinário e respiratório, além de sepse. Essas infecções são causadas por linhagens bacterianas geralmente multirresistentes. Genes que codificam as enzimas modificadoras de aminoglicosídeos (EMAs) e metiltransferases 16S RNAr podem estar presentes em isolados de enterobactérias também produtores de Klebsiella pneumoniae carbapapenemase (KPC). Portanto, o objetivo deste estudo foi investigar genes que codificam resistência aos aminoglicosídeos em 30 isolados de E. aerogenes e em 28 isolados de K. pneumoniae portadores do gene blaKPC resistentes a amicacina, tobramicina e/ou gentamicina, oriundos de colonização e infecção em pacientes de diferentes hospitais em Recife-PE, Brasil. A investigação dos genes armA, rmtB, rmtD, aac(3)Ia, aac(3)IIa, aac(6´)Ib, ant(2´)Ia e aph(3’)-VI foi realizada através de PCR, seguida de sequenciamento de DNA. Nos isolados de K. pneumoniae observou-se uma maior ocorrência dos genes ant(2´)Ia, seguidos de aac(3)IIa, aph(3’)-VI e aac(6´)Ib. O gene mais encontrado em E. aerogenes foi o aph(3’)-VI, seguidos de aac(3)-IIa e ant(2”)-Ia. Esse é o primeiro relato de aph(3’)-VI em E. aerogenes no Brasil. Os genes aac(3)-Ia, armA, rmtB e rmtD não foram encontrados. Esses achados ressaltam para a gravidade da alta ocorrência de isolados de K. pneumoniae e E. aerogenes portadores de genes para EMAs e gene blaKPC principalmente colonizando pacientes, visto que essas bactérias podem atuar na disseminação de mecanismos de resistência dentro da unidade hospitalar e limitar as opções de tratamento. / Klebsiella pneumoniae and Enterobacter aerogenes have been highlighted as important agents of healthcare-associated infections (HAIs), primarily causing wound, urinary and respiratory tracts infections, and sepsis. These infections are often caused by multiresistant bacterial strains. Genes encoding aminoglycoside modifying enzymes (AMEs) and 16S RNAr methyltransferases can also be present in Enterobacteriaceae isolates producing Klebsiella pneumoniae carbapapenemase (KPC). Therefore, the aim of this study was to investigate genes encoding resistance to aminoglycosides in 30 isolates of E. aerogenes and 28 K. pneumoniae isolates carrying the blaKPC gene and resistant to amikacin, tobramycin and / or gentamicin, from colonization and infection in patients from different hospitals in Recife-PE, Brazil. The investigation of the genes armA, rmtB, rmtD, aac(3)Ia, aac(3)IIa, aac(6´)Ib, ant(2´)Ia e aph(3’)-VI was performed by PCR followed DNA sequencing. In K. pneumoniae isolates there was a higher incidence of genes ant(2´)Ia, followed by aac(3)IIa, aph(3’)-VI e aac(6´)Ib. The gene most frequently found in E. aerogenes was aph(3’)-VI, followed by aac(3)-IIa e ant(2”)-Ia . This is the first report of aph (3 ') - VI in E. aerogenes in Brazil. The genes aac(3)-Ia, armA, rmtB e rmtD were not found. These findings points to the seriousness of the high incidence of isolates of K. pneumoniae and E. aerogenes carriers of AMEs and blaKPC gene, mainly colonizing patients, since these bacteria can act in the dissemination of resistance mechanisms within the hospital and to limit treatment options.

Dans la nature, les micro-organismes sont organisés en communautés agrégées dénommées biofilms, particulièrement adaptées à la survie en milieu hostile. Les difficultés pour prévenir la formation ou éliminer des biofilms matures par des stratégies conventionnelles ont encouragé le développement de nouvelles approches inspirées des mécanismes de compétition entre différents micro-organismes au sein de biofilms naturels. Au cours de ce travail, nous nous sommes intéressés à l'effet anti-biofilm de bactéries bénéfiques appartenant aux genres Lactobacillus et Bifidobacterium. Dans un premier temps, nous avons testé l'effet anti-biofilm de surnageants neutralisés vis-à-vis de deux pathogènes Klebsiella pneumoniae et Staphylococcus epidermidis dans un modèle expérimental statique. Si les extraits des quelques souches de Bifidobacterium testées stimulaient la formation de biofilm par K. pneumoniae sur surface abiotique, la majorité de ceux des 140 souches de Lactobacillus exerçait un effet inhibiteur et nous avons retenu une des souches dont le surnageant de culture entraînait une inhibition majeure (70%), Lactobacillus plantarum CIRM653. Cet extrait s'est également avéré capable de disperser des biofilms préformés à K. pneumoniae sur surface abiotique mais aussi d’inhiber la formation de biofilms sur surface biotique, et ce indépendamment d’un effet bactéricide. La formation de biofilms mixtes formés par L. plantarum et K. pneumoniae dans des modèles expérimentaux cinétiques a permis, comparativement à l'observation de biofilms mono-espèce à K. pneumoniae, de mettre en évidence des défauts de structuration du biofilm associés à une diminution de la biomasse de K. pneumoniae et une augmentation de celle de L. plantarum. Grâce à une approche transcriptionnelle ciblée, nous avons montré que L. plantarum induisait, par le biais de son surnageant, des modifications de l’expression de gènes impliqués dans la formation de biofilm chez K. pneumoniae. Quatre gènes impliqués dans le quorum-sensing (opérons lsr) étaient sous-exprimés et trois gènes de structure du pilus de type 3 étaient sur-exprimés. L'augmentation de la production de pili de type 3 fonctionnels a été validée par Western-blot et des tests d’hémagglutination. Cette surexpression est probablement responsable du niveau élevé des capacités d’adhésion sur surface abiotique d'agrégats de K. pneumoniae issus de la dispersion induite par L. plantarum.Le comportement des deux souches a également été testé in vivo, dans un modèle murin de colonisation intestinale par K. pneumoniae avec administration orale quotidienne de L. plantarum. Le dénombrement du pathogène dans les selles des animaux a montré qu'en présence de L. plantarum, K. pneumoniae maintient des niveaux de colonisation élevés, contrairement au contrôle (sans Lactobacillus) où une diminution graduelle est observée.Enfin, nous avons initié le développement d'un modèle expérimental tripartite permettant d'associer les deux partenaires bactériens avec des cellules épithéliales dans un système en flux continu. La réponse spécifique des cellules eucaryotes a également été abordée : nous avons pu mettre en évidence que L. plantarum exerçait un effet inhibiteur vis-à-vis de la réponse inflammatoire épithéliale pulmonaire induite par K. pneumoniae. En conclusion, la description d'une activité anti-biofilm in vitro ne serait pas synonyme d'une réduction in vivo de la colonisation de surfaces biotiques, mais à une plus grande capacité de dissémination. Ces observations démontrent l’importance d’une expertise précise de l’action des bactéries bénéfiques et de la maitrise du ratio bénéfice-risque pour leur utilisation. / In the natural environment microorganisms are organized in aggregated communities called biofilms, which are particularly adapted to the survival in harsh conditions. The difficulties to prevent the formation or elimination of mature biofilms by conventional strategies have encouraged the development of new approaches inspired by competition mechanisms occurring between microorganisms within natural biofilms.In this work, we looked for anti-biofilm effects of beneficial bacteria belonging to Lactobacillus and Bifidobacterium genus. We first tested the anti-biofilm effect of neutralized supernatants against both pathogens Klebsiella pneumoniae and Staphylococcus epidermidis in a static experimental model. The few Bifidobacterium extracts tested led to an increase in biofilm formation by K. pneumoniae on abiotic surface, whereas the majority of the 140 strains of Lactobacillus exerted an inhibitory effect. Lactobacillus plantarum CIRM653 was selected for further experiments because its culture supernatant displayed major inhibition (70%). This extract was also capable of dispersing preformed biofilms of K. pneumoniae on abiotic surface, but also able to inhibit biofilm formation on biotic surface, independently of a bactericidal effect. The formation of mixed biofilm containing L. plantarum and K. pneumoniae in kinetic experimental models highlighted the biofilm structure defects associated with a decrease of K. pneumoniae biomass and an increase of that of L. plantarum, compared to a monospecies K. pneumoniae biofilm. Targeted transcriptional approach was used to assess changes in the expression of genes involved in biofilm formation by K. pneumoniae after contact with L. plantarum supernatant. Four genes involved in quorum-sensing (operons lsr) were under-expressed and three type 3 pili structural genes were over-expressed. The increase of functional surface located type 3 pili was validated by Western blotting and hemagglutination tests. This overexpression was probably responsible for the observed high level of adhesion capacity to abiotic surfaces of K. pneumoniae aggregates recovered after dispersion induced by L. plantarum.The behavior of the two strains was also tested in vivo in a K. pneumoniae murine intestinal colonization model with daily oral administration of L. plantarum. Viable cells counting of the pathogen in the animals’ feces showed that K. pneumoniae maintained high levels of colonization in the presence of L. plantarum, unlike the control (without Lactobacillus) where a gradual decrease was observed.Finally, we initiated the development of a tripartite experimental model allowing the combination of the two bacterial partners with epithelial cells in a continuous flow system. In parallel, the specific response of eukaryotic cells to these bacteria was addressed: L. plantarum exerted an inhibitory effect on the pulmonary epithelial inflammatory response induced by K. pneumoniae.In conclusion, these results highlight the discrepancy between in vitro anti-biofilm activity of L. plantarum and its in vivo behavior leading to increased dissemination of the pathogen. Substantial expertise of beneficial bacteria is therefore necessary to fully assess their benefit-risk ratio.