Influence of environmental factors on adhesion of yeasts to abiotic surfaces

Adhezija i formiranje biofilma kvasaca na kontaktne površine 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 šteti zdravlju potrošača. 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 površine je ograničeno na samo neke vrste roda Candida, a naročito malo se zna o vrstama roda Pichia kao adhezivnim agensima. Cilj istraživanja u okviru ove doktorske disertacije je bio da se proceni potencijal adhezije sojeva kvasaca Candida i Pichia na polistiren površinu, diskove od nerđajućeg čelika (AISI 304, Ra = 25.20 – 961.9 nm) i drveta sa različitim stepenom hrapavosti površine, i ispita uticaj hrapavosti površine na adheziju kvasaca. Hidrofobnost površine ćelije (CSH) sojeva iz rodova Candida i Pichia je određena u cilju procene korelacije između hidrofobnosti površine 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 površine diskova od nerđajućeg čelika. Rezultati su pokazali da su svi testirani sojevi kvasaca Candida i Pichia su bili u stanju da adheriraju na polistiren, površine 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, što 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, što ukazuje da hidrofobnost površine 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 površine nerđajućeg čelika, dok hrapavost drvenih površina 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 površinama 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. Naši podaci takođe ukazuju da je temperatura veoma važan faktor koji značajno utiče na adheziju ovih kvasaca. Problem adhezije i formiranje biofilma kvasaca na abiotičkim površinama 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 što 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.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’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. 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