Infection of the human urinary tract is one of the commonest bacterial infections, with uropathogenic E.coli (UPEC) being responsible for 90% of the diagnosed cases, with significant morbidity and mortality. The urinary bladder is a remarkable autonomic musculomembranous organ under conscious control. Its two main functions are, storage and voiding of urine. Any disturbance to normal urination leads to various clinical conditions, such as urinary incontinence, bladder retention, overactive bladder syndrome, prostatitis in men and urinary tract infections (UTI). Determining the predisposition of an individual to UTI by discovering a biomarker would allow for a more rational selection of patients who might best benefit from either antibiotic prophylaxis or preemptive surgical intervention. The purpose of this study was to examine the epigenetic effects of UPEC infection directly, or indirectly in the bladder. The study also identified potential gene candidates, such as TLR4 and CTCF, for development of DNA methylation biomarker targets.

Uropathogenic E. coli (UPEC) is the most frequent cause of urinary tract infection (UTI), being responsible for up to 85% of community acquired and 40% of nosocomial cases. UPEC strains harbour various virulence factors that contribute to their ability to cause disease. The high prevalence across the globe of multidrug resistant UPEC is a significant threat to therapy. Virulent and resistant UPEC strains have been recognised as belonging to major lineages and we have only recently begun to understand the factors contributing to their successful global dissemination. Work in this thesis was carried out to identify the population structure of E. coli isolates recovered from urosepsis and biliary sepsis, to reveal any differences in genetic background. A total of 100 isolates from the blood and urine of 50 patients presenting with urosepsis and 27 isolates from cases of biliary sepsis were subjected to genotypic and phenotypic analysis, including MLST, virulence gene detection and antibiogram and metabolic profiling. Urosepsis paired isolates showed identical genotypes and antimicrobial resistance profiles. However, several pairs of isolates showed discrepant metabolic activity profiles suggesting niche specific regulation of metabolism. Members of the ST131 clone were significantly associated with antibiotic resistance and ST38 isolates were associated with the highest level of metabolic activity. An in vivo infection model was used to investigate the virulence potential of isolates from the major UPEC lineages. Galleria mellonella larvae inoculated with ST69 and ST127 isolates showed significantly higher mortality rates than those infected with other strains. However, one isolate of ST127 (strain EC18) was avirulent and comparative genomic analyses with a single virulent ST127 strain revealed an IS1 mediated deletion in the O-antigen cluster in strain EC18, which is likely to explain the lack of virulence in the larvae and demonstrates the importance of this cell surface molecule in the model system. Finally, a total of 202 UPEC isolates were recovered from community and hospital urine samples from a tertiary care hospital in Riyadh, Saudi Arabia. Molecular epidemiological investigation of the strains was carried out to examine the overall UPEC population structure, for the first time in any part of Saudi Arabia. The most common lineages were ST131 (17.3%), ST73 (11.4%), ST38 (7.4%), ST69 (7.4%) and ST10 (6.4%). The findings highlight the successful spread of multidrug resistant, CTX-M positive ST38, ST131 and ST405 UPEC in Saudi Arabia. The high proportion (35%) of ESBL producing E. coli isolates is a particular concern and is driving frequent prescription of carbapenem antibiotics. A total of four isolates of ST38 were positive for aggR, which is a virulence marker of enteroaggregative E. coli (EAEC); ST38 strains that cause UTI but have an EAEC genetic background are becoming recognised as novel UPEC and this clonal group warrants further study.

Orientador: Rodrigo Tavanelli Hernandes / Resumo: Escherichia coli uropatogênica (UPEC) causa a maioria das infecções do trato urinário (ITU), incluindo cistite e pielonefrite, no hospedeiro humano. A UPEC utiliza numerosos fatores de virulência para entrar, aderir, colonizar, adquirir nutrientes essenciais, multiplicar e causar danos ao ambiente do trato urinário. Estudos recentes demonstraram que alguns isolados de UPEC carregam fatores de virulência associados à patótipos diarreiogênicos de E. coli (DEC), como EAEC (E. coli enteroagregativa) e EPEC (E. coli enteropatogênica). Uma grande preocupação nas infecções por UPEC é o aumento da resistência antimicrobiana, levando à falha do tratamento em algumas ITUs causadas por esse patógeno. Nesse estudo, um total de 118 isolados de UPEC de amostras ambulatoriais de urina de pacientes atendidos no Hospital das Clinicas da Faculdade de medicina de Botucatu entre março e maio de 2018. Reação em cadeia da polimerase (PCR) foi usada para detectar 29 genes que codificam fatores de virulência, bem como marcadores de DEC (escN, stx1/2, aatA e aggR); além de genes que codificam adesinas e toxinas associadas ao patótipo EAEC. Os isolados de UPEC foram designados nos diferentes filogrupos de E. coli, utilizando um PCR quadruplex; e a determinação do perfil de susceptibilidade antimicrobiana foi realizada pelo método de disco difusão. Entre os isolados estudados, 39,8% foram atribuídos ao filogrupo B2, enquanto UPEC dos filogrupos B1 (14,4%), A (14,4%), D (12,7%), F (8,5%), G (3,4%), E ( ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Uropathogenic Escherichia coli (UPEC) cause the majority of urinary tract infections (UTIs), including cystitis and pyelonephritis, in the human host. UPEC utilizes numerous virulence factors to entry, adhere, colonize, acquire essential nutrients, multiply and cause damage in the urinary tract environment. Recent studies have shown that some UPEC isolates carry virulence factors associated with the diarrheagenic E. coli (DEC) pathotypes, such as EAEC (enteroaggregative E. coli) and EPEC (enteropathogenic E. coli). A major concern in UPEC infections is the constant increasing of antimicrobial resistance, thus leading to treatment failure in some UTIs caused by this pathogen. In this study a total of 118 UPEC isolates were obtained from outpatient urine samples, attended at University Hospital of Botucatu Medical School between March and May of 2018. Polymerase chain reaction (PCR) was used to detect 29 virulence factor-encoding genes, diarhoeagenic E. coli markers, (escN, stx1/2, aatA and aggR), as well as genes encoding adhesins and toxins associated with the EAEC pathotype. The UPEC isolates were assigned in the distinct E. coli phylogroups, using a quadruplex PCR; and the determination of the antimicrobial resistance profile was performed using the diskdiffusion method. Among the isolates studied, 39.8% were assigned to phylogroup B2, while UPEC isolates from other phylogroups were detected as follows: B1 (14,4%), A (14,4%), D (12,7%), F (8,5%), G (3,4%), E ( 2,5%), E. cla... (Complete abstract click electronic access below) / Mestre

Yes / Infection by pathogenic bacteria on implanted and indwelling medical devices during surgery causes large morbidity and mortality worldwide. Attempts to ameliorate this important medical issue have included development of antimicrobial surfaces on materials, “no touch” surgical procedures, and development of materials with inherent low pathogen attachment. The search for new materials is increasingly being carried out by high throughput methods. Efficient methods for extracting knowledge from these large data sets are essential. Data from a large polymer microarray exposed to three clinical pathogens is used to derive robust and predictive machine-learning models of pathogen attachment. The models can predict pathogen attachment for the polymer library quantitatively. The models also successfully predict pathogen attachment for a second-generation library, and identify polymer surface chemistries that enhance or diminish pathogen attachment. / CSIRO Advanced Materials Transformational Capability Platform. Newton Turner Award for Exceptional Senior Scientists. Wellcome Trust. Grant Number: 085245. NIH. Grant Number: R01 DE016516

As ITU são as mais freqüentes infecções ocasionadas por ExPEC. Entre os fatores de virulência (FV) encontram-se nestas cepas adesinas, invasinas, toxinas, sideróforos, e evasinas, localizados em plasmídios ou ilhas de patogenecidade. O objetivo deste estudo foi caracterizar 45 cepas de E. coli isoladas de 33 cães e 7 gatos com ITU, quanto aos sorotipos, FV e grupos filogenéticos. Dos sorogrupos relacionados às ITU foram encontrados O6 (20%), O2 (16%), O25 (4%), O4 e O11 (4% cada um). Entre os genes pesquisados, foram encontrados fimH (100%), pap (47%), sfa (33%) e iha (4%); ibeA (29%); cnf1 (31%), hlyA (27%); fyuA (80%), iucD (22%); traT (51%); cvaC (20%) e malX (67%). Os isolados felinos foram agrupados em B2 (89%) e D (11%), enquanto os caninos em A (5,5%), B1 (19,5%), B2 (55,5%) e D (19,5%). Estes resultados sugerem que as ExPEC isoladas de cães e gatos apresentam potencial patogênico para ocasionar doenças mais graves que as ITU, assim como ocorre em humanos. Além disso, a similitude com as amostras humanas reforça a hipótese acerca de seu potencial zoonótico. / The ability of ExPEC to cause extraintestinal infections in humans, dogs, and cats is associated with the expression of a variety of virulence factors (VF). The aim of this study was to evaluate the frequency of VF related to ExPEC, serotypes, and phylogenetic groups in 45 strains isolated from 33 dogs and 7 cats with UTI. These strains presented serogroups related with extraintestinal infections, e.g. O6 (20%), O2 (16%), O25 (4%), O4 e O11 (each one) and the following genes: fimH (100%), pap (47%), sfa (33%) e iha (4%); ibeA (29%); cnf1 (31%), hlyA (27%); fyuA (80%), iucD (22%); traT (51%); cvaC (20%) e malX (67%), cvaC (20%), and malX (67%). All feline strains were concentrated in B2 (89%) and D (11%) phylogenetic groups, whereas the canine ones were distributed in the four groups, A (5,5%), B1 (19,5%), B2 (55,5%) and D (19,5%). These findings suggesting that ExPEC isolated from dog and cat contain virulence markers to cause diseases, more severe than UTI, likewise in humans. Besides, these the close similarity between human and animal ExPEC supports the hypotesis of zoonotic potencial of them.

As ITU são as mais freqüentes infecções ocasionadas por ExPEC. Entre os fatores de virulência (FV) encontram-se nestas cepas adesinas, invasinas, toxinas, sideróforos, e evasinas, localizados em plasmídios ou ilhas de patogenecidade. O objetivo deste estudo foi caracterizar 45 cepas de E. coli isoladas de 33 cães e 7 gatos com ITU, quanto aos sorotipos, FV e grupos filogenéticos. Dos sorogrupos relacionados às ITU foram encontrados O6 (20%), O2 (16%), O25 (4%), O4 e O11 (4% cada um). Entre os genes pesquisados, foram encontrados fimH (100%), pap (47%), sfa (33%) e iha (4%); ibeA (29%); cnf1 (31%), hlyA (27%); fyuA (80%), iucD (22%); traT (51%); cvaC (20%) e malX (67%). Os isolados felinos foram agrupados em B2 (89%) e D (11%), enquanto os caninos em A (5,5%), B1 (19,5%), B2 (55,5%) e D (19,5%). Estes resultados sugerem que as ExPEC isoladas de cães e gatos apresentam potencial patogênico para ocasionar doenças mais graves que as ITU, assim como ocorre em humanos. Além disso, a similitude com as amostras humanas reforça a hipótese acerca de seu potencial zoonótico. / The ability of ExPEC to cause extraintestinal infections in humans, dogs, and cats is associated with the expression of a variety of virulence factors (VF). The aim of this study was to evaluate the frequency of VF related to ExPEC, serotypes, and phylogenetic groups in 45 strains isolated from 33 dogs and 7 cats with UTI. These strains presented serogroups related with extraintestinal infections, e.g. O6 (20%), O2 (16%), O25 (4%), O4 e O11 (each one) and the following genes: fimH (100%), pap (47%), sfa (33%) e iha (4%); ibeA (29%); cnf1 (31%), hlyA (27%); fyuA (80%), iucD (22%); traT (51%); cvaC (20%) e malX (67%), cvaC (20%), and malX (67%). All feline strains were concentrated in B2 (89%) and D (11%) phylogenetic groups, whereas the canine ones were distributed in the four groups, A (5,5%), B1 (19,5%), B2 (55,5%) and D (19,5%). These findings suggesting that ExPEC isolated from dog and cat contain virulence markers to cause diseases, more severe than UTI, likewise in humans. Besides, these the close similarity between human and animal ExPEC supports the hypotesis of zoonotic potencial of them.