<p>Adhezija i formiranje biofilma kvasaca na kontaktne povr&scaron;ine materijala je ozbiljan higijenski problem u prehrambenoj industriji, jer može biti izvor kontaminacije tokom proizvodnje, dovodeći do velikih ekonomskih gubitaka zbog kvarenja hrane i smanjenja kvaliteta proizvoda, a takođe može da &scaron;teti zdravlju potro&scaron;ača. S drge strane, mnoge infekcije u bolnici uzrokovane su adhezijom vrsta roda Candida na medicinska sredstva i implantate. Međutim, trenutno razumevanje adhezije i kolonizacije kvasaca na abiotičke povr&scaron;ine je ograničeno na samo neke vrste roda Candida, a naročito malo se zna o vrstama roda Pichia kao adhezivnim agensima.<br />Cilj istraživanja u okviru ove doktorske disertacije je bio da se proceni potencijal adhezije sojeva kvasaca Candida i Pichia na polistiren povr&scaron;inu, diskove od nerđajućeg čelika (AISI 304, Ra = 25.20 &ndash; 961.9 nm) i drveta sa različitim stepenom hrapavosti povr&scaron;ine, i ispita uticaj hrapavosti povr&scaron;ine na adheziju kvasaca. Hidrofobnost povr&scaron;ine ćelije (CSH) sojeva iz rodova Candida i Pichia je određena u cilju procene korelacije između hidrofobnosti povr&scaron;ine ćelije i adhezije kvasaca na polistiren. Pored toga, ispitan je uticaj različitih medijuma, temperatura, i različitih biljnih ekstrakata na adheziju odabranih sojeva iz rodova Candida i Pichia na povr&scaron;ine diskova od nerđajućeg čelika. Rezultati su pokazali da su svi testirani sojevi kvasaca Candida i Pichia su bili u stanju da adheriraju na polistiren, povr&scaron;ine od nerđajućeg čelika i drveta, ali u različitoj meri. Soj vrste C. albicans je imao veću sposobnost adhezije na polistiren i nerđajući čelik u odnosu na sojeve vrsta C. glabrata, C. parapsilosis i C. krusei. U slučaju kod roda Pichia, sojevi vrste P. membranifaciens su pokazali manju sposobnost adhezije na nerđajući čelik nego ispitivani soj vrste P. pijperi, &scaron;to nije slučaj kod adhezije na polistiren. Na osnovu dobijenih rezultata pokazano je da su sojevi vrsta C. glabrata, C. parapsilosis, C. krusei, P. pijperi i P. membranifaciens bili izrazito do umereno hidrofobni, dok je soj vrste C. albicans bio hidrofilan, &scaron;to ukazuje da hidrofobnost povr&scaron;ine ćelije (CSH) nije značajno u korelaciji sa količinom ćelija adheriranih na polistiren. Pored toga, na adheziju sojeva kvasaca Candida i Pichia značajno je uticala hrapavost povr&scaron;ine nerđajućeg čelika, dok hrapavost drvenih povr&scaron;ina nije značajno uticala. Dalje mi smo ispitali uticaj različitih medijuma za rast i temperature na adheziju odabranih kvasaca. Primećeno je da je količina adheriranih ćelija kvasaca C. albicans i C. glabrata na povr&scaron;inama od nerđajućeg čelika bila znatno veća u Malt Extract Broth (MEB) medijumu, dok je za C. parapsilosis, C. krusei, P. pijperi i P. membranifaciens bila u Yeast Peptone Dextrose (YPD) medijumu. Na&scaron;i podaci takođe ukazuju da je temperatura veoma važan faktor koji značajno utiče na adheziju ovih kvasaca. Problem adhezije i formiranje biofilma kvasaca na abiotičkim povr&scaron;inama i rastuća otpornost na postojeće antimikrobne agense predstavilo je izazov za nas za pronalaženjem novih alternativnih agenasa biljnog porekla. Stoga mi smo ispitali antimikrobnu i antibiofilm aktivnost biljnih ekstrakata kao &scaron;to su Humulus lupulus, Alpinia katsumadai i Evodia rutaecarpa prema C. albicans, C. glabrata i P. membranifaciens. Na osnovu dobijenih MIC vrednosti, testirani biljni ekstrakati su bili efikasni u inhibiciji kvasaca. Primećeno je da su biofilmovi C. glabrata pokazali veću otpornost na biljne ekstrakte u odnosu na C. albicans. Međutim, ekstrakti A. katsumadai i E. rutaecarpa su promovisani rast i razvoj formiranog biofilma P. membranifaciens.</p> / <p>The adhesion and biofilm formation of yeast on the contact surfaces of the material is a serious hygiene problem in the food industry, because it can be a source of contamination during production, leading to major economic losses due to food spoilage and reduced product quality, and can also affects the consumer&rsquo;s health. On the other hand, many infections in the hospital are caused by the adhesion of Candida spp. to medical devices and implants. However, the understanding of adhesion and colonization of yeasts on abiotic surfaces is limited to only some Candida species, and in particular little is known about Pichia species as adhesive agents.<br />The aim of the research in this doctoral dissertation was to assess the potential of Candida and Pichia strains to adhere to polystyrene surface, stainless steel discs (AISI 304, Ra = 25.20 - 961.9 nm) and wood (beech) with different degrees of surface roughness, and examined the influence of surface roughness on yeast adhesion. Cell surface hydrophobicity (CSH) of Candida and Pichia strains was determined in order to assess the correlation between the cell surface hydrophobicity and the yeast adhesion to polystyrene. In addition, the influence of different medium, temperature, and various plant extracts on the adhesion of selected Candida and Pichia strains to to stainless steel surfaces was examined. Our results indicated that all Candida and Pichia strains were able to adhere to polystyrene, stainless steel surfaces and wood in a species- and strain-dependent manner. C. albicans had a higher ability to adhere to polystyrene and stainless steel compared to C. glabrata, C. parapsilosis and C. krusei. Regarding Pichia species, P. membranifaciens strains were less adherent to stainless steel than P. pijperi, what is not the case in the adhesion to polystyrene. Based on the results obtained, C. glabrata, C. parapsilosis, C. krusei, P. pijperi i P. membranifaciens strains were strongly to moderately hydrophobic while only C. albicans was moderately hydrophilic, indicating that the cell surface hydrophobicity is not significantly correlated with the amount of cells adhered to polystyrene. In addition, the adhesion of Candida and Pichia strains was significantly influenced by the roughness of stainless steel surface, while the roughness of wood surfaces did not significantly influence. Further on we examined the impact of different growth medium and temperature on adhesion of selected yeasts. It was noted that the amount of adhered cells of C. albicans and C. glabrata on stainless steel surfaces were significantly higher in the Malt Extract Broth (MEB) medium, whereas for C. parapsilosis, C. krusei, P. pijperi and P. membranifaciens it was Yeast Peptone Dextrose (YPD) medium. Our data also indicated that temperature is a very important factor which considerably affects the adhesion of these yeast. The problem of yeast adhesion and biofilm formation on abiotic surfaces and its growing resistance to existing antimicrobial agents, presented a challenge for us to find new alternative agents of plant origin. Hence, we investigated antimicrobial and antibiofilm activity of plant extracts such as Humulus lupulus, Alpinia katsumadai and Evodia rutaecarpa against C. albicans, C. glabrata and P. membranifaciens. According to the MIC values, all plant extracts were effective in the inhibition of yeast strains. It was observed that biofilms of C. glabrata are more resistance to plant extracts as compared to C. albicans. However, extracts of A. katsumadai and E. rutaecarpa were promoted the growth and development of a preformed biofilm of P. membranifaciens.</p>

Made available in DSpace on 2016-01-26T18:56:03Z (GMT). No. of bitstreams: 1 Deigilam Cestari Esteves.pdf: 2776358 bytes, checksum: 7e1a46b5b264cbf8e0c0d96e68b501e2 (MD5) Previous issue date: 2014-12-18 / The environmental and hospital infections, caused by bacteria which are resistant to a wide variety of antibiotics, have shown increasing records in the last years, manifesting themselves in high mortality and lethality. Recent researches report that the bacteria presents a survival profile on dry surfaces in order to maintain their virulence when exposed to biological fluids such as urine, saliva and blood. The objective of this study was to document - through laboratorial analysis the survival capacity of the main bacteria of medical interest on abiotic surfaces. The adopted procedures were completely conducted in the microbiology laboratory at Unoeste, in Presidente Prudente SP. Standard ATCC strains of Staphylococcus aureus (ATCC25923), Enterococcus faecalis (ATCC29212), Escherichia coli (ATCC35218), Klebsiella pneumoniae (ATCC700603) and Pseudomonas aeruginosa (ATCC27583) were used, to which biological fluids and water were added. The surfaces were tile, synthetic fabric, mattress and cotton fabric. The surfaces were contaminated with suspension composed by the ATCC strains, biological fluids and water in addition to the control containing only the ATCC strains, which were stored in petri dishes, kept in room temperature. Every seven days the surfaces were dipped in trypticase soy broth and kept in the oven for 24h at 37ºC. They were spread from the contaminated brothin Mueller Hinton agar and kept for 24h in the oven at 37ºC. The viability analysis was done through the colony-forming unity (CFU) counting. For the statistical analysis, the Software R was used, through the Friedman and Kruskal Wallis tests for multiple comparisons, analyzing within each factor (solution or surface) which differ between one another (Friedman s test) and also which treatments interactions are different (Kruskal-Wallis). Through this analysis it was possible to observe that in the current work that Staphylococcus aureus kept its viability for a longer period than other microorganisms under all the tested conditions, presenting significant differences between the fluids and surfaces, with a particular colony growth in blood and cotton. The Klebsiella pneumoniae showed differences between fluids, with saliva containing the highest colony number. With regards to the other tested bacteria, there was no isolated significance. For the multiple comparisons analysis, only the Pseudomonas aeruginosa presented no significant difference between any pair of treatments, whereas the other bacteria presented significant differences between the correlations. The need to analyze the environmental impact of these microorganisms persistence in environments, which are vulnerable to human beings, guides the creation of measures in order to control the spread of pathogenic microorganisms. / As infecções ambientais e hospitalares causadas por bactérias resistentes a um amplo espectro de antibióticos têm índices crescentes nos últimos anos, manifestando-se com alta morbidade e letalidade. Pesquisas recentes evidenciam que as bactérias demonstram um perfil de sobrevivência, em superfícies secas de modo a manter sua virulência quando expostas a fluidos biológicos como urina, saliva e sangue. O objetivo desse estudo foi documentar através de análises laboratoriais a capacidade de sobrevivência das principais bactérias de interesse médico em superfícies abióticas. Os procedimentos foram totalmente realizados no laboratório de microbiologia da Unoeste em Presidente Prudente - SP., utilizando cepas padrão ATCC de Staphylococcus aureus(ATCC25923), Enterococcus faecalis (ATCC29212), Escherichia coli(ATCC35218), Klebsiella pneumoniae(ATCC700603) e Pseudomonas aeruginosa(ATCC27583), ao qual foram adicionados fluídos biológicos e água. As superfícies utilizadas foram piso, tecido sintético, colchão e tecido algodão. As superfícies foram contaminadas com suspensão composta pelas cepas ATCC, fluídos biológicos e água, além do controle contendo somente as cepas ATCC, armazenadas em placas de petri e mantidas em temperatura ambiente. A cada sete dias as superfíciesarmazenadas foram mergulhadas em caldo Tripscaseína de Soja (TSB), colocadas na estufa por 24h a 37ºC. Foram semeados do caldo contaminado em ágar Mueller Hinton e mantidos por 24h em aquecimento a 37º em Estufa.A análise da viabilidadefoi realizada através da contagem de unidade formadora de colônias (U.F.C.). Para a análise estatística utilizou o software R, realizando os testes de Friedman e de Kruskal Wallis para comparações multiplas, analisando dentro de cada fator (solução ou superfície) quais diferem entre si (Teste de Friedman) e, também, quais interações de tratamentos são diferentes (Kruskal-Wallis). Através dessas análises foi possível observar no presente trabalho que Staphylococcus aureus manteve a viabilidade por tempo maior que os outros microrganismos em todas as condições testadas, apresentando diferença significativa entre os fluídos e as superfícies, tendo sangue e tecido algodão crescimento de maior número de colônias. A Klebsiella pneumoniae apresentou diferença entre os fluídos, sendo asaliva com maior número de colônias. Para as outras bactérias testadas não houve significância isoladamente. Para as análises de comparações múltiplas somente para a Pseudomonas aeruginosanão houve diferença significativa entre algum par de tratamentos, as outras bactérias apresentaram diferenças significativas entre as correlações. A necessidade de analisar o impacto ambiental da persistência desses microrganismos em ambientes vulneráveis ao ser humano, norteia o delineamento de medidas para o controle na disseminação de microrganismos patogênicos.

Made available in DSpace on 2016-07-18T17:46:20Z (GMT). No. of bitstreams: 1 Deigilam Cestari Esteves.pdf: 2776358 bytes, checksum: 7e1a46b5b264cbf8e0c0d96e68b501e2 (MD5) Previous issue date: 2014-12-18 / The environmental and hospital infections, caused by bacteria which are resistant to a wide variety of antibiotics, have shown increasing records in the last years, manifesting themselves in high mortality and lethality. Recent researches report that the bacteria presents a survival profile on dry surfaces in order to maintain their virulence when exposed to biological fluids such as urine, saliva and blood. The objective of this study was to document - through laboratorial analysis the survival capacity of the main bacteria of medical interest on abiotic surfaces. The adopted procedures were completely conducted in the microbiology laboratory at Unoeste, in Presidente Prudente SP. Standard ATCC strains of Staphylococcus aureus (ATCC25923), Enterococcus faecalis (ATCC29212), Escherichia coli (ATCC35218), Klebsiella pneumoniae (ATCC700603) and Pseudomonas aeruginosa (ATCC27583) were used, to which biological fluids and water were added. The surfaces were tile, synthetic fabric, mattress and cotton fabric. The surfaces were contaminated with suspension composed by the ATCC strains, biological fluids and water in addition to the control containing only the ATCC strains, which were stored in petri dishes, kept in room temperature. Every seven days the surfaces were dipped in trypticase soy broth and kept in the oven for 24h at 37ºC. They were spread from the contaminated brothin Mueller Hinton agar and kept for 24h in the oven at 37ºC. The viability analysis was done through the colony-forming unity (CFU) counting. For the statistical analysis, the Software R was used, through the Friedman and Kruskal Wallis tests for multiple comparisons, analyzing within each factor (solution or surface) which differ between one another (Friedman s test) and also which treatments interactions are different (Kruskal-Wallis). Through this analysis it was possible to observe that in the current work that Staphylococcus aureus kept its viability for a longer period than other microorganisms under all the tested conditions, presenting significant differences between the fluids and surfaces, with a particular colony growth in blood and cotton. The Klebsiella pneumoniae showed differences between fluids, with saliva containing the highest colony number. With regards to the other tested bacteria, there was no isolated significance. For the multiple comparisons analysis, only the Pseudomonas aeruginosa presented no significant difference between any pair of treatments, whereas the other bacteria presented significant differences between the correlations. The need to analyze the environmental impact of these microorganisms persistence in environments, which are vulnerable to human beings, guides the creation of measures in order to control the spread of pathogenic microorganisms. / As infecções ambientais e hospitalares causadas por bactérias resistentes a um amplo espectro de antibióticos têm índices crescentes nos últimos anos, manifestando-se com alta morbidade e letalidade. Pesquisas recentes evidenciam que as bactérias demonstram um perfil de sobrevivência, em superfícies secas de modo a manter sua virulência quando expostas a fluidos biológicos como urina, saliva e sangue. O objetivo desse estudo foi documentar através de análises laboratoriais a capacidade de sobrevivência das principais bactérias de interesse médico em superfícies abióticas. Os procedimentos foram totalmente realizados no laboratório de microbiologia da Unoeste em Presidente Prudente - SP., utilizando cepas padrão ATCC de Staphylococcus aureus(ATCC25923), Enterococcus faecalis (ATCC29212), Escherichia coli(ATCC35218), Klebsiella pneumoniae(ATCC700603) e Pseudomonas aeruginosa(ATCC27583), ao qual foram adicionados fluídos biológicos e água. As superfícies utilizadas foram piso, tecido sintético, colchão e tecido algodão. As superfícies foram contaminadas com suspensão composta pelas cepas ATCC, fluídos biológicos e água, além do controle contendo somente as cepas ATCC, armazenadas em placas de petri e mantidas em temperatura ambiente. A cada sete dias as superfíciesarmazenadas foram mergulhadas em caldo Tripscaseína de Soja (TSB), colocadas na estufa por 24h a 37ºC. Foram semeados do caldo contaminado em ágar Mueller Hinton e mantidos por 24h em aquecimento a 37º em Estufa.A análise da viabilidadefoi realizada através da contagem de unidade formadora de colônias (U.F.C.). Para a análise estatística utilizou o software R, realizando os testes de Friedman e de Kruskal Wallis para comparações multiplas, analisando dentro de cada fator (solução ou superfície) quais diferem entre si (Teste de Friedman) e, também, quais interações de tratamentos são diferentes (Kruskal-Wallis). Através dessas análises foi possível observar no presente trabalho que Staphylococcus aureus manteve a viabilidade por tempo maior que os outros microrganismos em todas as condições testadas, apresentando diferença significativa entre os fluídos e as superfícies, tendo sangue e tecido algodão crescimento de maior número de colônias. A Klebsiella pneumoniae apresentou diferença entre os fluídos, sendo asaliva com maior número de colônias. Para as outras bactérias testadas não houve significância isoladamente. Para as análises de comparações múltiplas somente para a Pseudomonas aeruginosanão houve diferença significativa entre algum par de tratamentos, as outras bactérias apresentaram diferenças significativas entre as correlações. A necessidade de analisar o impacto ambiental da persistência desses microrganismos em ambientes vulneráveis ao ser humano, norteia o delineamento de medidas para o controle na disseminação de microrganismos patogênicos.