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Traballo Fin de Grao en Medicina. Curso 2020-2021 Los antibióticos se consideran uno de los grandes éxitos de la medicina del siglo XX. Sin embargo, su uso inadecuado y abuso han hecho que en la actualidad nos encontremos con un problema con graves consecuencias: la resistencia bacteriana a los antibióticos. Este, ha pasado de ser considerado un problema individual a un problema global y de salud pública. Además, en los últimos años, empezó a abordarse desde un punto de vista ecológico y no solo desde el punto de vista sanitario como se estaba haciendo hasta entonces al descubrirse que, el desarrollo de bacterias resistentes no solo se relaciona con el uso de antibióticos sino también con cualquier alteración del equilibrio natural. Prueba de ello es la influencia que tienen los genes bacterianos responsables de la resistencia a metales pesados, en el desarrollo y evolución de las resistencias a antibióticos por su proximidad a los genes que median las mismas. Recientes estudios demuestran que las Estaciones Depuradoras de Aguas Residuales (EDARs) desempeñan un papel importante en la propagación y difusión de resistencias bacterianas al medioambiente. Varios de estos trabajos constatan un incremento de los genes de resistencia a antibióticos y de bacterias resistentes a los mismos en medios acuáticos como resultado de un marcado aumento del consumo de antibióticos por parte de la población a nivel mundial. Se han detectado bacterias resistentes a antibióticos en aguas residuales urbanas, líquidos residuales hospitalarios, lodos cloacales, aguas subterráneas y ríos contaminados por el vertido de aguas residuales. Este trabajo consiste en una revisión sistemática. En él se aportará información sobre los principales mecanismos que hacen a las bacterias resistentes a los antibióticos y, sobre el papel que desempeñan las aguas residuales mediante los procesos biológicos, físicos y químicos que tienen lugar en el proceso de depuración de estas, en la propagación de las resistencias Os antibióticos considéranse uns dos grandes éxitos da medicina do século XX. Porén, o seu uso inadecuado e o seu abuso fixeron que na actualidade nos atopemos cun problema con graves consecuencias: a resistencia bacteriana aos antibióticos. Este, pasou de ser considerado un problema individual a un problema global e de saúde pública. Ademais, nos últimos anos, comezou a abordarse dende un punto de vista ecolóxico e non só dende un punto de vista sanitario como se estaba facendo ata entón, ao descubrirse que, o desenvolvemento de bacterias resistentes non só se relaciona co uso de antibióticos senón tamén con calquera alteración do equilibrio natural. Proba disto é a influencia que teñen os xenes bacterianos responsábeis da resistencia a metais pesados, no desenvolvemento e evolución das resistencias aos antibióticos pola súa proximidade aos xenes que median as mesmas. Recentes estudos demostran que as Estacións Depuradoras de Augas Residuais (EDARs) desempeñan un papel importante na propagación e difusión de resistencias bacterianas ao medioambiente. Varios destes traballos constatan un incremento dos xenes de resistencia a antibióticos e de bacterias resistentes aos mesmos en medios acuáticos como resultado dun marcado aumento do consumo de antibióticos por parte da poboación a nivel mundial. Detectáronse bacterias resistentes a antibióticos en augas residuais urbanas, líquidos residuais hospitalarios, lodos cloacais, augas subterráneas e ríos contaminados polo vertido de augas residuais. Este traballo consiste nunha revisión sistemática. Nel aportarase información sobre os principais mecanismos que fan ás bacterias resistentes aos antibióticos e sobre o papel que desenvolven as aguas residuais mediante os procesos biolóxicos, físicos e químicos que teñen lugar no proceso de depuración das mesmas, na propagación das resistencias Antibiotics are considered one of the great successes of twentieth-century medicine. However, their inappropriate use and abuse have led to a problem with serious consequences: bacterial resistance to antibiotics. This has gone from being considered an individual problem to a global and public health problem. In addition, in recent years, it began to be approached from an ecological point of view and not only from a sanitary point of view, as was being done until then when it was discovered that the development of resistant bacteria is not only related to the use of antibiotics but also with any alteration of the natural balance. Proof of this is the influence of the bacterial genes responsible for resistance to heavy metals in the development and evolution of antibiotic resistance due to their proximity to the genes that mediate them. Recent studies show that Wastewater Treatment Plants (WWTPs) play an important role in the propagation and diffusion of bacterial resistance to the environment. Several works confirm an increase in antibiotic resistance genes and antibiotic resistant bacteria in aquatic environments as a result of a marked increase in the consumption of antibiotics by the world population. Antibiotic resistant bacteria have been detected in urban wastewater, hospital waste liquids, sewage sludge, groundwater and rivers contaminated by wastewater discharge. This work consists of a systematic review. It will provide information on the main mechanisms that make bacteria resistant to antibiotics and on the role that wastewater plays through the biological, physical and chemical processes that take place in the process of purifying them, in the propagation of the resistors

Dissertação de mestrado em Técnicas de Caracterização e Análise Química, Uma ferida cutânea se não for devidamente tratada pode incorrer numa ferida crónica suscetível a infeções generalizadas, as quais podem resultar na morte do paciente. Atualmente, a engenharia de tecidos é descrita como um campo interdisciplinar que combina princípios de engenharia, química e biologia para gerar soluções que permitam reparar, restaurar e/ou melhorar as funções de tecidos lesados. No mesmo sentido, a área têxtil aborda soluções com base em fibras poliméricas, produzidas a partir de uma vasta gama de polímeros, que permitem gerar estruturas com uma grande área superficial, porosidade e resistência mecânica passivas de serem utilizadas como pensos bioativos que promovam uma atividade cicatrizadora e antimicrobiana eficiente. Os péptidos com atividade antimicrobiana (AMPs) têm despertado muita atenção pela possibilidade de serem uma alternativa promissora aos antibióticos, na busca de agentes mais eficazes contra as bactérias, evitando que estas desenvolvam resistência e por ser possível a sua síntese e manipulação para serem aplicados como o elemento biomédico nos pensos bioativos. Este projeto resulta da colaboração entre o Departamento de Química e o Centro de Ciência e Tecnologia Têxtil (2C2T), e visa a síntese dos AMPs Peptaibolin e Tiger 17, através de metodologias de síntese em fase sólida assistida por micro-ondas, a sua caracterização química, estrutural e antimicrobiana, e posterior imobilização em estruturas nanofibrosas poliméricas produzidas por eletrospinning. Para atingir este objetivo, técnicas como a Ressonância magnética nuclear de protão (1 HRMN) e a Cromatografia líquida de alta eficiência (HPLC) foram aplicadas na caraterização do péptido, assim como testes antimicrobianos de concentração mínima inibitória ou cinética de evolução microbiana. A técnica de eletrospinning foi utilizada para o processamento de nanofibras a partir do polímero natural, acetato de celulose (CA), e do sintético, policaprolactona (PCL). As matrizes poliméricas geradas foram caracterizadas recorrendo a técnicas como MO, SEM, ATR-FTIR, TGA, DSC, ensaios de hidratação e determinação indireta de porosidade, e medições de ângulos de contacto e, consequente, determinação da energia de superfície. Sendo possível concretizar todos os objetivos propostos para este trabalho, e fruto de uma extensa otimização de processos e investigação futura, esta inovação poderá resultar numa terapia única com vista a proteger as feridas contra infeções, e simultaneamente acelerar a cicatrização e regeneração dos tecidos., A skin wound if not properly treated can result in a chronic wound susceptible to widespread infections, which can result in the patient's death. Currently, tissue engineering is described as an interdisciplinary field that combines principles of engineering, chemistry and biology to generate solutions that allow to repair, restore and/or improve the functions of injured tissues. In the same sense, the textile area addresses solutions based on polymeric fibers, produced from a wide range of polymers, which allow the generation of structures with a large surface area, porosity and mechanical resistance that can be used as bioactive dressings that promote a healing and efficient antimicrobial activity. Antimicrobial peptides (AMPs) have attracted a lot of attention due to the possibility of being an alternative to antibiotics in the search for more effective agents against bacteria, preventing them from developing resistance. They can be applied as the biomedical element in bioactive dressings because their synthesis and manipulation are possible. This project results from the collaboration between the Department of Chemistry and the Center for Science and Textile Technology (2C2T), and aims at the synthesis of AMPs Peptaibolin and Tiger 17, through solid phase synthesis methodologies assisted by microwave, their chemical, structural and antimicrobial characterization, and subsequent immobilization on polymeric nanofiber structures produced by electrospinning. To achieve this objective, techniques such as Proton Nuclear Magnetic Resonance (1 H RMN) and High Performance Liquid Chromatography (HPLC) were applied to the peptide characterization, as well as antimicrobial tests with minimal inhibitory concentration or kinetics of microbial evolution. The electrospinning technique was used to process nanofibers from the natural polymer, cellulose acetate (CA), and the synthetic, polycaprolactone (PCL). The polymeric matrices generated were characterized using techniques such as MO, SEM, ATR-FTIR, TGA, DSC, hydration tests and indirect porosity determination, measurements of contact angles and, consequently, determination of surface energy. As it is possible to achieve all the objectives proposed for this work, and as a result of extensive optimization process and future research, this innovation may result in a unique therapy with a view to protecting wounds against infections, while simultaneously accelerating healing and tissue regeneration., Gostava ainda de agradecer ao Centro de Ciência e Tecnologia Têxtil (2C2T) por permitir a realização deste projeto nos seus laboratórios e ainda à Fundação da Ciência e Tecnologia (FCT) pelo financiamento do projeto onde este trabalho foi inserido, projeto PEPTEX PTDC/CTM-TEX/28074/2017 (POCI-01-0145-FEDER-028074)

To tackle one of the biggest global health problems, the resistance of microorganisms to antibiotics, a collective effort in the search for more effective agents against bacteria was required. Peptides with antimicrobial activity have been raising much attention as a promising alternative for antibiotics. Peptaibols, for instance, are a family of antimicrobial peptides (AMPs) with great biomedical potential, in which the Peptaibolin can be highlighted. This peptide has gained relevance due to its small amino acids content, only four, and its acetyl group and a phenylalaninol residue (Phol) at the N-terminal and C-terminal, respectively. Here, we report the synthesis of Peptaibolin through Solid Phase Peptide Synthesis assisted by Microwave heating (MW-SPPS) in a pre-loaded Phe-Wang resin. Starting from a loading of 0.51 mmol/g, two syntheses were made, using two different combinations of coupling reagents. The best option was DIC/Oxima, achieving a yield of 50.0%. Proton Nuclear Magnetic Resonance (1H-NMR) studies confirmed the peptide structure, while High Performance Liquid Chromatography (HPLC) verified the peptide purity. The peptide solubility was examined against several combinations of solvents. Peptaibolin was not soluble in water, only in organic solvents or in the combination of both. Antimicrobial testing was conducted using Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, and Pseudomonas aeruginosa. Minimum inhibitory concentration studies demonstrated the resistance of bacteria to the peptide action and the peptide instability in bacterial growth conditions, Fundação para a Ciência e Tecnologia-FCT (Portugal) for funding through CQUM (UID/QUI/00686/2020) and project PTDC/QUI-COL/28052/2017. The NMR spectrometer Bruker Avance III 400 is part of the National NMR Network and was purchased within the framework of the National Program for Scientific Re-equipment, contract REDE/1517/RMN/2005 with funds from POCI 2010 (FEDER) and FCT. Authors also acknowledge FCT for funding the project PEPTEX with reference PTDC/CTM-TEX/28074/2017 (POCI-01-0145-FEDER-028074). They acknowledge project UID/CTM/00264/2021 of Centre for Textile Science and Technology (2C2T), funded by national funds through FCT/MCTES

Background Urinary tract infections constitute a major public health concern. The aim of the study is to look into the antibiotic sensitivity profile of uropathogenic bacteria among diabetic individuals in the Diabetology Unit of the Bafoussam Regional Hospital, West Cameroon. Methods A cross-sectional study was carried out in Bafoussam Regional Hospital. Urine was collected in a sterile jar previously labelled. The microorganisms were isolated on agar medium and their final identification was carried out on the API20E gallery. The antibiogram was performed using agar diffusion methods. Results Escherichia coli (25.30%) and Staphylococcus aureus (19.27%) were the most noticed species in the diabetic patients, whereas Escherichia coli (32.00%) and Klebsiella pneumoniae (22.00%) were the most noticed in non-diabetic patients. There was a significant association between antibiotic resistance and diabetic status (for ceftriaxone: X2 = 23.78 and P-value < 0.001; for cefixime: X2 = 19.31 and P-value < 0.001; for ceftazidime: X2 = 9.45 and P-value = 0.008; for cefotaxime: X2 = 10.97 and P-value = 0.004; for cefepime: X2 = 27.93 and P-value < 0.001; and for ciprofloxacin: X2 = 11.13 and P-value = 0.003). Multidrug resistance rate against some bacterial species were higher in diabetic patients (62.50% for Escherichia coli, 63.16% for Klebsiella pneumoniae, and 78.57% for Staphylococcus aureus) compared with non-diabetic patients (37.50% for Escherichia coli, 36.84% for Klebsiella pneumoniae, and 21.43% for Staphylococcus aureus). Conclusions This study revealed that there is an association between antibiotic resistance and diabetic status. Research and interventions must be focused on the elderly diabetic population in order to fight against the occurrence of drug-resistant uropathogenes.

L’organisation mondiale de la santé estime que le choléra entraîne 100, 000 décès par an pour environ 4 millions de cas recensés, plaçant ainsi cette maladie comme un enjeu de santé publique majeur. Cette infection est causée par Vibrio cholerae, une bactérie à Gram négatif vivant en milieu aquatique. Face à cette agression, l’épithélium intestinal et les bactéries du microbiote agissent comme une barrière, en exprimant notamment des peptides antimicrobiens (PAM). V. cholerae, comme de nombreux pathogènes, montre une résistance accrue aux PAM. Malgré une avancée constante sur la compréhension des mécanismes de résistances bactériens aux PAM, de nombreuses inconnues demeurent, principalement en raison des techniques de mutagénèse aléatoire utilisées pour leur identification. L’objectif de cette étude est d’identifier de nouveaux mécanismes de résistance impliqués dans la résistance de V. cholerae aux PAM. Grâce à des expériences de séquençage par spectrométrie masse suivi d’études bio-informatiques, nous avons identifié les protéines OmpV et Lap comme des candidates intéressantes pour être impliquées dans la résistance de V. cholerae. Alors que la souche V. cholerae A1552 délétée du gène ompV (DompV) ne semble pas présenter de défaut de croissance ou de perméabilité, nos résultats ont montré une diminution des concentrations minimales inhibitrices en différents PAM tels que LL-37. Les tests fonctionnels semblent suggérer l’implication des vésicules de sécrétion dans le mécanisme de d’OmpV face à LL-37 chez V. cholerae., According to the World Health Organization, cholera remains a significant health problem, causing 100,000 death per 4 millions infections per year. V. cholerae, the etiologic agent of cholera and a Gram-negative bacterium, is generally transmitted via contaminated food or water. During infection, the microbiota and intestinal epithelium acts as a barrier by producing antimicrobial peptides (AMP). Resistance to AMP has emerged as a virulence factor in pathogens and specifically in V. cholerae. As AMP are considered as novel molecular therapeutic agents, it is truly important to better understand strategies of resistance of V. cholerae. The aim of this study is to identify unknown mechanisms involved in V. cholerae resistance to AMP. For this purpose, after protein sequencing by mass spectrometry (MS-MS) and bioinformatics, we identified the OmpV porin, member of the outer membrane protein family and the Lap protease as new protein candidates for strategies of resistance. We characterized the V. cholerae A1552 strain deleted for ompV (DompV). We confirmed OmpV implication in AMP resistance by minimum inhibitory concentration and did not observed any differences in growth or permeability in the presence of AMP between the wild-type (wt) and the (DompV) strains. The work presented here suggest an implication of outer membrane vesicules in V. cholerae resistance mechanisms to some AMP as LL-37.

U današnje vrijeme velika je prisutnost i upotreba antibiotika u liječenju. Oni se propisuju svakodnevno na bolničkoj i lokalnoj razini od strane liječnika opće medicine. Česta je pojava prekomjernog liječenja antibioticima, a posljedica je problematika rezistencije, odnosno otpornost bakterija i bakterijskih bolesti na antibiotske lijekove. Značajan broj pacijenata nesavjesno pristupa liječenju antibioticima ne pridržavajući se uputa liječenja, ne poštujući vremenski rok liječenja, a mnogi pacijenti štede antibiotik u svrhu kasnijeg samoliječenja. Edukacija o antibiotskom liječenju od strane medicinskog tima je vrlo važan korak za smanjenje bespotrebne i nedisciplinirane konzumacije antibiotika. Veliki broj oboljelih vjeruje da antibiotici djeluju na viruse, a ne na bakterijske infekcije. S obzirom na problematiku liječenja antibioticima, provedeno je istraživanje kojemu je cilj bilo ispitati znanje i stavove opće populacije o antibioticima i o važnosti pravilne primjene peroralnih antibiotika u svrhu dobivanja jasnije slike stvarne problematike. U istraživanju su sudjelovali ispitanici svih životnih dobi, različitih stupnjeva obrazovanja, ispitanici s i bez medicinske struke kako bi se rezultatima dobio širok spektar mišljenja. Istraživanje je provedeno online anketom na društvenim mrežama putem Google obrasca u periodu od 20.6.2018. do 20.8.2018. Anketa je podijeljena na dva dijela. U prvom dijelu prikupljali su se opći podaci o ispitaniku, a drugi dio sastojao se od 15 pitanja o općem znanju te korištenju antibiotskih lijekova. Istraživanje je provedeno na uzorku od 181 ispitanika. Nowadays there is an increasing presence and use of antibiotics in the treatment. Antibiotics are prescribed daily to at hospital level, and local by general practitioners. Consequently an increased presence of antibiotics can lead to the excessive antibiotic treatment that results in the problems of resistance, or resistance of bacteria and bacterial infections to antibiotic drugs. With the patients in turn leads to the fact that a significant number of patients careless approach to treatment with antibiotics is not following the instructions of treatment, disregarding the time limit treatment, wrong care or saving antibiotics for the purpose of later self-healing, etc. Education of antimicrobial therapy by the medical team is a very important step for reducing unnecessary and undisciplined consumption of antibiotics. Although we live in a time when our health information available at any time, either through the media and the Internet, either through books, magazines, scientific articles, a large number of patients believe that antibiotics work on viruses, but not to a bacterial infection. Given the situation and the problem of antibiotic treatment in the health survey was conducted which was aimed to examine the knowledge and attitudes of the general population on antibiotics and see the views and opinions of the respondents on the importance of the proper implementation of oral antibiotics in order to obtain a clearer picture of the actual problems. In a study involving participants of all ages, different grade levels, subjects with and without the medical profession in order to obtain the results of a wide range of opinions. The survey was conducted on-line survey on social networks through Google form over a period of 20.6.2018.- 20.8.2018. The survey is divided into two parts. In the first part were collected general information about the subject, and the second part consisted of 15 questions on general knowledge and the use of antibiotic drugs. The study was conducted on the sample of 181.

ÖZET BANKNOTLARDA GRAM NEGATİF BAKTERİ VARLIĞI VE ANTİBİYOTİK DİRENCİNİN ARAŞTIRILMASITürkiye OECD ülkeleri arasında antibiyotik kullanımının en fazla olduğu ülkedir. Dolayısıyla antibiyotik direnci konusunu sadece hastane ortamından izole edilen bakterilerde değil toplumda ve çevrede de araştırmak epidemiyolojik veri eldesi ve dirençle mücadelede yeni yaklaşımların belirlenmesi bakımından anlamlı olacaktır. Toplumda dirençli bakterilerin yayılımı konusunda yaygın dolaşımda bulunan materyal olan banknotlar, konunun araştırılması bakımından uygun görülmüş; çalışmamızda dirençli bakteri sorununda en büyük paya sahip Gram negatif basiller başta olmak üzere dirençli bakterilerin banknotlarda varlığını araştırmak amaçlanmıştır. Banknotların, dirençli bakterilerin taşınmasında bir risk faktörü olup olmadığı yorumlanmıştır.İstanbul'da Aralık 2018- Ocak 2019 tarihlerinde farklı bölgelerden 5 TL ve 10 TL'lik toplam 200 adet banknot toplanmış olup kültür, tanımlama ve antibiyotik duyarlılık testleri gerçekleştirilmiştir. 5 TL'lik banknotlarda saptanan bakteri sayısı 36, 10 TL'lik banknotlarda ise 22 olarak bulunmuştur. Tespit edilen bakterilerin %60'ı 5 TL'lik banknotlarda görülmüştür. Staphylococcus aureus %12,2, Enterococcus spp. %14, Escherichia coli %24, Klebsiella pneumoniae %22,8, Pseudomonas aeuroginosa %8,77, Serratia marcescens %3,5, Kluyvera cryocrescens %1,75 ve Salmonella spp. %3,5 oranında bulunmuştur. Dirençli bakteriler olarak metisiline dirençli Staphylococcus aureus (MRSA) %5,2, Vankomisine dirençli Enterococcus spp. (VRE) %7,01 ve genişlemiş spektrumlu beta laktamaz (ESBL) yapan Enterobacteriaceae üyeleri %17,4 oranında bulunmuştur. Çalışmamızda banknotların dirençli bakterilerle kontamine olduğu gösterilmiş olup dirençli bakterilerin taşınmasında bir risk faktörü olabileceği sonucuna ulaşılmıştır.Anahtar Kelimeler: Banknot, Gram Negatif Bakteri, Gram Pozitif Bakteri, Antibiyotiklere Direnç Turkey is the country where antibiotic usage is most common among OECD countries. Therefore, it would be meaningful to investigate the issue of antibiotic resistance not only in bacteria isolated from the hospital environment but also in the community and the environment in terms of obtaining epidemiological data and identifying new approaches in the fight against resistance.Banknotes, which are widely circulating material in the society which makes appropriate subject to investige the role on spreading antibiotic resistant . In our study, we aimed to investigate the presence of resistant bacteria in banknotes, especially Gram negative bacilli, which have the largest share in the problem of resistant bacteria. It was interpreted whether banknotes were a risk factor for the transport of resistant bacteria.A total of 200 banknotes of 5TL and 10TL were collected from different regions in Istanbul between December 2018 and January 2019 and culture, identification and antibiotic susceptibility tests were performed. The number of bacteria were isoleted from 5-note and 10-note banknotes were found to be 36 and 22 respectively. Which were defined as Staphyloococcus aureus (12.2%), Enterococcus spp. (14%), E.coli (24%), Klebsiella pneumoniae (22.8%), Pseudomonas aeuroginosa (8.77%), Serratia marcescens (3.5%), Kluyvera cryocrescens (1.75%) and Salmonella spp. It was found to be (3.5%). Methicillin-resistant staphylococcus aureus (MRSA) was resistant to (5.2%) and vancomycin-resistant enterococcus spp. (VRE) 7.01% and extended spectrum beta lactase (ESBL) were found in (17,4 %).In our study, it was shown that banknotes were contaminated with resistant bacteria and it was concluded that it may be a risk factor in the transportion of resistant bacteria among community.Keywords: Paper Money, Gram Negative Bacteria, Gram Positive Bacteria, Bacteria Resistance 74

Biofilms give to bacteria micro-environmental benefits; confers protection against antimicrobials. Bacteria have antibiotic resistance by conventional and unusual mechanisms leading to delayed wound healing, to increase recurrent chronic infections and nosocomial contamination of medical devices. Objective: This narrative review aims to introduce the characteristics of Bacteria-biofilms, antimicrobial resistance mechanisms and potential alternatives for prevention and control of its formation. Methods: Search strategy was performed on records: PubMed / Medline, Lilacs, Redalyc; with suppliers such as EBSCO and thesaurus MeSH and DeCS. Conclusions: Knowledge and research performance of biofilm bacteria are relevant in the search of technology for detection and measuring sensitivity to antibiotics. The identification of Bacterial-biofilms needs no-traditional microbiological diagnosis., Los biofilms otorgan a las bacterias que las producen, ventajas micro-medio ambientales significativas que les confiere además protección frente a los antimicrobianos. Las bacterias-biofilms hacen resistencia a los antibióticos por mecanismos convencionales e inusuales, conducen al retardo en la cicatrización de heridas, al incremento de infecciones recurrentes, crónicas e intrahospitalarias por contaminación de dispositivos médicos. Objetivo: esta revisión narrativa tiene como propósito dar a conocer las características de las bacterias-biofilms, sus mecanismos de resistencia antimicrobiana y las potenciales alternativas de prevención y control. Métodos: revisión narrativa. La estrategia de búsqueda se realizó en PubMed/ Medline, Lilacs y Redalyc con los proveedores EBSCO. Se empleó los tesauros MeSH y DeCS. Conclusiones: es importante la necesidad de profundizar en el conocimiento e investigación del funcionamiento de las bacterias-biofilm, en la búsqueda de tecnología para su detección y medición de la sensibilidad frente a los antibióticos, teniendo en cuenta que la identificación de estas asociaciones microbianas se aleja del diagnóstico microbiológico tradicional.

International audience; BackgroundLow-dose azithromycin has beneficial effects on severity of the lung disease in cystic fibrosis (CF) patients for a period of 6–12 months after initiation of the treatment. Although its impact in the longer term is uncertain, this treatment is frequently used chronically. The aim of this retrospective study was to investigate the effects of low-dose azithromycin treatment on the progression of CF lung disease in patients treated for more than 12 months.MethodsAll of the CF patients followed in our pediatric center and who had been on low-dose azithromycin for more than 12 sequential months were included. The clinical data were collected for one year before and three years after the initiation of the azithromycin treatment. These data comprised lung function analyses, rates of exacerbations and of antibiotic courses, and changes in the airways' bacterial colonization.ResultsA total of 68 patients were included (mean age: 9.95 yrs (3.61)). After 12 months, significant reductions in the numbers of pulmonary exacerbations and antibiotic courses were present. However, this effect was not maintained in the subsequent periods, during which increased rates of both pulmonary exacerbations and antibiotic courses were observed. The lung function decline was not modified during the treatment, and a decreasing time-dependent trend typical of CF was observed for the various parameters. No differences in the airway colonization by pathogens such as P. aeruginosa and methicillin-sensitive and/or -resistant Staphyloccocus aureus were observed during the treatment. However, isolated Staphyloccocus aureus strains became resistant to macrolides after 6 months of azithromycin and remained resistant thereafter.ConclusionsNo clinical benefits of low-doses azithromycin were present after one year of treatment in young CF patients. Selection for macrolide-resistant strains of bacteria occurred, which should lead to a reconsideration of the duration of azithromycin treatment in CF.

Antibiotics are among the most commonly prescribed drugs, however optimal dosages are not yet well defined. The aim of this thesis was to develop pharmacokinetic (PK) and pharmacokinetic-pharmacodynamics (PK/PD) models that characterize the course of antimicrobial drug concentrations and effects over time, with an emphasis on the development of resistance. These models were applied to optimize dosing regimens of antimicrobial therapies.A population PK model for colistin and its prodrug, colistin methanesulfonate (CMS) was developed in critically ill patients receiving colistin by nebulization and/or undergoing an intermittent hemodialysis (HD). Results predicted clear benefits of using aerosol delivery of 2MIU CMS dose for the treatment of pulmonary infections. For patients with HD session dosing regimen of CMS should be 1.5 MIU twice daily with an additional dose of 1.5 MIU after each HD session.An assessment of the performances of different PK-PD models by using a simulation approach have shown the importance of longer study designs and of complementary microbiological data to predict accurately bacterial resistance development. A semi-mechanistic PK/PD model that incorporates mutation rate and adaptive resistance development of a bioluminescent strain of Pseudomonas aeruginosa against colistin wasdeveloped based on in-vitro data. A high, quick and partially reversible resistance was described. These results confirm that the first 24 h of treatment are critical in the management of infections, that colistin alone cannot eradicate completely the mutants of Pseudomonas aeruginosa that were selected during the experiments and that combination therapies seem necessary.; Les antibiotiques sont actuellement parmi les médicaments les plus utilisés, mais les schémas thérapeutiques optimaux ne sont pas toujours bien définis. Le but de cette thèse était de développer des modèles pharmacocinétiques (PK) et pharmacodynamiques (PK/PD) décrivant les profils de concentrations des antibiotiques ainsi que leurs effets et le développement de résistances bactériennes afin d'optimiser les schémas thérapeutiques.Un modèle PK de population sur la colistine et sa prodrogue, le colistine methanesulfonate (CMS), a été développé chez les patients recevant la colistine par voie aérosol et/ou sous hémodialyse (HD). Les résultats ont montré un net avantage de la voie aérosol pour le traitement des infections pulmonaires avec une dose de 2 MUI de CMS. Pour les patients sous HD une dose de 1.5 MUI de CMS 2 fois par jour est recommandée avec une dose supplémentaire de 1.5 MUI de CMS après chaque séance de HD.L'évaluation des performances de différents modèles PK/PD via à une approche par simulation a montré l'importance d'effectuer des études suffisamment longues ainsi que d'obtenir des données microbiologiques complémentaires afin de décrire le développement de la résistance bactérienne.Un modèle PK/PD incluant taux de mutation et résistance adaptative à la colistine d'une souche bioluminescente de Pseudomonas aeruginosa a été développé à partir de données in-vitro. Une résistance rapide, importante et partiellement réversible a été décrite. Ces résultats confirment l'importance des 24 premières heures dans le traitement des infections, que la colistine seule ne peut pas complètement éliminer les mutants de Pseudomonas aeruginosa et que des associations semblent nécessaires.

Antibiotics are among the most commonly prescribed drugs, however optimal dosages are not yet well defined. The aim of this thesis was to develop pharmacokinetic (PK) and pharmacokinetic-pharmacodynamics (PK/PD) models that characterize the course of antimicrobial drug concentrations and effects over time, with an emphasis on the development of resistance. These models were applied to optimize dosing regimens of antimicrobial therapies.A population PK model for colistin and its prodrug, colistin methanesulfonate (CMS) was developed in critically ill patients receiving colistin by nebulization and/or undergoing an intermittent hemodialysis (HD). Results predicted clear benefits of using aerosol delivery of 2MIU CMS dose for the treatment of pulmonary infections. For patients with HD session dosing regimen of CMS should be 1.5 MIU twice daily with an additional dose of 1.5 MIU after each HD session.An assessment of the performances of different PK-PD models by using a simulation approach have shown the importance of longer study designs and of complementary microbiological data to predict accurately bacterial resistance development. A semi-mechanistic PK/PD model that incorporates mutation rate and adaptive resistance development of a bioluminescent strain of Pseudomonas aeruginosa against colistin wasdeveloped based on in-vitro data. A high, quick and partially reversible resistance was described. These results confirm that the first 24 h of treatment are critical in the management of infections, that colistin alone cannot eradicate completely the mutants of Pseudomonas aeruginosa that were selected during the experiments and that combination therapies seem necessary., Les antibiotiques sont actuellement parmi les médicaments les plus utilisés, mais les schémas thérapeutiques optimaux ne sont pas toujours bien définis. Le but de cette thèse était de développer des modèles pharmacocinétiques (PK) et pharmacodynamiques (PK/PD) décrivant les profils de concentrations des antibiotiques ainsi que leurs effets et le développement de résistances bactériennes afin d'optimiser les schémas thérapeutiques.Un modèle PK de population sur la colistine et sa prodrogue, le colistine methanesulfonate (CMS), a été développé chez les patients recevant la colistine par voie aérosol et/ou sous hémodialyse (HD). Les résultats ont montré un net avantage de la voie aérosol pour le traitement des infections pulmonaires avec une dose de 2 MUI de CMS. Pour les patients sous HD une dose de 1.5 MUI de CMS 2 fois par jour est recommandée avec une dose supplémentaire de 1.5 MUI de CMS après chaque séance de HD.L'évaluation des performances de différents modèles PK/PD via à une approche par simulation a montré l'importance d'effectuer des études suffisamment longues ainsi que d'obtenir des données microbiologiques complémentaires afin de décrire le développement de la résistance bactérienne.Un modèle PK/PD incluant taux de mutation et résistance adaptative à la colistine d'une souche bioluminescente de Pseudomonas aeruginosa a été développé à partir de données in-vitro. Une résistance rapide, importante et partiellement réversible a été décrite. Ces résultats confirment l'importance des 24 premières heures dans le traitement des infections, que la colistine seule ne peut pas complètement éliminer les mutants de Pseudomonas aeruginosa et que des associations semblent nécessaires.

Since the 1940s, antibiotics have greatly reduced the adverse effects of infectious diseases caused by microbes. However, due to excessive, and often incorrect, use of known antibiotics, many organisms have adapted antibiotic resistance. Currently, over 70% of known infectious bacteria are resistant to at least one antibiotic. In the U.S. , 90,000 deaths occur each year due to infection by bacteria resistant to antibiotics. This number has increased by nearly 75,000 in the last 20 years. It is necessary, therefore, to continue developing new antibiotics in an effort to keep up with increasing antibiotic resistance. Traditional in vitro and whole cell drug screens are inefficient. Unlike in vitro studies, in vivo studies can rule out toxic compounds early in analysis. They can also test compounds in the presence of important host/pathogen relationships. The action of an antimicrobial can therefore be viewed within the context of the whole organism, and not just at the drug-receptor site, as in vitro . Current in vivo testing is done in small mammals, such as mice. Mice, however, are resource intensive to maintain. Often, the antibiotic compounds being tested are toxic to the mice, killing them and creating a need for new miceB��� Our lab group has proposed the use of the nematode Caenorhabditis e/egans for in vivo tests as an intermediate step prior to testing in mammals.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior Poultry production is a very important economic activity in Brazil once the country is among the highest world avian producers. The knowledge of the pathogens epidemiology is essential to the control of infectious diseases and such control is very strict in chickens and turkeys on the avian industry. However, the avian population of the country shows a big diversity of domestic and wild birds and the chicken backyard population is out of this control. To investigate the presence of antibodies against specific viruses in the backyard chicken population, blood was collected of 867 non-vaccinated birds, from 60 farms and 22 counties from the Rio Grande do Sul State, Brazil. Neutralizing antibodies against infectious bronchitis virus (IBV) were detected in 65% (564/867) of the individuals, against avian reovirus (ARV) in 21.6% (187/867) and, against infectious bursal disease virus (IBDV) in 80.2% (695/867). The results presented on the first chapter indicated that the tested viruses are circulating among this population. Among the wild species there is a group of Galliformes, classified at the Cracidae family and commonly known as guans, chachalacas and curassows which deserved a special attention in the present study. To determine the microbiota, the bacterial resistance and the presence of antibodies against specific viruses in these birds, fifty one serum and cloacal swab samples were collected from 10 different cracid species captive in the Rio Grande do Sul State during 2007. Serum samples were submitted to serum-neutralization test and specific antibodies were detected against IBV in 5.9% (3/51) of the cracids, against ARV in 15.7% (8/51) and, against IBDV in 35.3% (18/51). All the samples were found to be negative to fowlpox virus by the AGID test. Bacterial isolation and identification were performed from the cloacal swabs. After that, the isolates were tested for antimicrobial susceptibility. Ninety three bacterial isolates were obtained from 10 different genera. Escherichia spp., Staphylococcus spp. and Streptococcus spp. are among the most prevalent genera. Antimicrobial resistance was observed in several isolates, with Serratia marcescens presenting the highest level of resistance to multi drugs followed by Streptococcus spp., Staphylococcus aureus and Escherichia coli, among others. The results from the second chapter of this dissertation allowed us to conclude that bacterial resistance is spread in the captive cracid microbiota and, most importantly, these species are susceptible to the infection by IBV, IBDV and ARV. A avicultura é uma das principais atividades econômicas do Brasil, que ocupa posição de destaque entre os exportadores de frango e subprodutos. O conhecimento da epidemiologia de patógenos que podem gerar prejuízos a essa atividade é essencial para o controle das enfermidades infecciosas. Este controle é realizado em larga escala em galinhas e perus criados no sistema industrial. Entretanto a população avícola do país consiste de uma grande diversidade de aves domésticas e selvagens, sendo o grupo conhecido como galinhas de terreiro formado por indivíduos desta espécie criados fora do sistema industrial. Com o objetivo de investigar a presença de anticorpos contra alguns vírus específicos na população de galinhas de terreiro, foram coletadas amostras de sangue de 867 galinhas não vacinadas, em 60 propriedades de 22 municípios do Estado do Rio Grande do Sul, Brasil, que foram testadas pela técnica de soroneutralização. Anticorpos contra o vírus da bronquite infecciosa das galinhas (IBV) foram detectados em 65% (564/867) destas aves, contra o reovírus aviário (ARV) em 21,6% (187/867) e contra o vírus da doença infecciosa da bolsa (IBDV) em 80,2% (695/867). Os descritos no primeiro capítulo desta dissertação permitiram demostrar a circulação dos vírus testados na população descrita. No segundo capítulo trabalhou-se com aves da ordem Galliformes, pertencentes à família Cracidae e conhecidos popularmente como jacus, jacutingas, araquãs e mutuns, com o intuito de conhecer a microbiota, resistência bacteriana e a presença de anticorpos contra vírus de aves, 51 amostras de soro e swab cloacal de 10 diferentes espécies de cracídeos cativos do Estado do Rio Grande do Sul foram coletadas durante o ano de 2007. As amostras de soro foram utilizadas para a detecção de anticorpos neutralizantes contra o IBV, ARV e IBDV, onde obteve os seguintes resultados: contra o IBV em 5,9% (3/51) das amostras positivas, contra o ARV em 15,7% (8/51) e contra o IBDV em 35,3% (18/51). Através do teste de imunodifusão em gel de ágar (IDGA) determinou-se que todas as amostras de soro eram negativas para o vírus da bouba aviária. Para o isolamento e identificação bacteriana foram realizados dos swabs cloacais e, posteriormente, foi testada a susceptibilidade antimicrobiana. Foram obtidos 93 isolados bacterianos, divididos em 10 diferentes gêneros. Entre os gêneros bacterianos mais prevalentes estavam Escherichia spp., Staphylococcus spp. e Streptococcus spp. Foi observada em grande número de isolados resistência à antimicrobianos, sendo que a bactéria Serratia marcescens apresentou o maior índice de resistência múltipla antimicrobiana. Os altos índices de resistência também foram detectados para Streptococcus spp., Staphylococcus aureus e Escherichia coli. Os resultados apresentados permitem concluir que a resistência bacteriana está disseminada na microbiota de cracídeos cativos e que indivíduos destas espécies são suscetíveis à infecção pelo IBV, IBDV e ARV, assim como as galinhas de terreiro.

Ficolins are soluble plasma and tissue proteins that are putatively involved in innate resistance of pigs to some bacteria containing N-acetylglucosamine in their surface polysaccharides. These studies were designed to determine how plasma ficolin concentration changes with age during the peri-weaning period and if porcine ficolins are induced in the liver during the acute phase response. Plasma ficolin concentrations of piglets weaned conventionally (28 days) or by segregated early weaning (14 days), were similar at 14 (25 ± 12[mu]g/ml), 35 (17 ± 11 [mu]g/ml) and 49 (26 ± 11 [mu]g/ml) days of age, using an indirect ELISA developed. In pigs exposed to ' E. coli' lipopolysaccharide, increased amounts of known porcine acute phase proteins, but not ficolins, were demonstrated by 2D SDS-PAGE of 35S-labeled proteins secreted by isolated hepatocytes. These studies demonstrate that plasma ficolins are constitutively expressed but are not induced during the acute phase response in young pigs.