Sezen BOLAT, Ayşe KALKANCI, Murat DİZBAY, Kayhan ÇAĞLAR, Işıl FİDAN, Semra KUŞTİMUR, ÖZLEM GÜZEL TUNÇCAN

Comparison of biomaterials in ability for gene expression levels of ALS1 and HWP1 in candida albicans and ICA1 in Staphylococcus epidermidis

Bu çalışma lateks-silikon kaplı, polivinil klorid, polivinilprolidon kaplı poliüretan ve poliüretan kateterlerin in-vitro biyofilm modelinde adezyon genlerinin ifadelenmesine etkilerini karşılaştırmak için düzenlenmiştir. Candida albicans’ın “aglutinin-like sequence” (ALS1), “hypha-spesific surface protein” (HWP1) genleri ve Staphylococcus epidermidis’in “intracellular adhesion” (ICA1) geni hedeşenmiştir. Dört farklı biyolojik madde ile dört farklı biyofilm modeli oluşturulmuştur. biyofilm katmanı kateter yüzeyinden sıyrılmış ve bu kitleden genomik mRNA izole edilerek gerçek zamanlı polimeraz zincir reaksiyonu (PCR) cihazında ölçülmüştür. Silikon kaplı idrar sondası yüzeyinde oluşan biyofilmde ALS1 ve HWP1 genlerinin kopya sayıları ortalaması, sırasıyla 2.50x105 ve 1.30x104 olarak, ICA geni kopya sayısı ortalama 1.80x105 kopya/ml olarak bulunmuştur. Yapısı polivinil klorid olan trakeal tüp yüzeyinde oluşan biyofilmde ALS1 ve HWP1 genlerinin kopya sayıları ortalaması, sırasıyla 7.56x106 ve 6.88x105 kopya/ml olarak, ICA gen kopya sayısı ortalaması ise 4.20x105 kopya/ml bulunmuştur. Polivinilprolidin kaplı poliüretan malzemeden yapılan nazogastrik tüp ve poliüretan malzemeden yapılan aspirasyon sondası yüzeyinde biyofilm oluşmadığı ve bu malzeme ile karşılaştırılan C.albicans örneğinde ALS1 ve HWP1 ile S.epidermidis örneğinde ICA gen ifedelerinin bulunmadığı gösterilmiştir. Polivinilprolidin kaplı poliüretan yüzeyde ALS 2.70x102, HWP1 2.27x101 ve ICA1 2.30x101 düzeyinde iken, poliüretan yüzeyinde ALS1, HWP1 ve ICA genleri sırası ile 1.50x101, 1.20x101 ve 3.10x101 bulunmuştur. Adezyon genleri polivinil klorid sonda yüzeyinde silikon olandan daha yüksek oranda gösterilmiştir. En düşük gen ifadelenmesi polivinilprolidon kaplı poliüretan kateter ve tek başına poliüretan yapılı kateterlerde gösterilmiştir. Kullanılan malzemelere bağlı olarak adezyon genlerinin arasında fark bulunması, klinik kullanımda bazı malzemelerin tercih edilmesini ve bazılarından sakınılmasını sağlayabilir.

Biyomaddelerin candida albicans ALS1 ve HWP1 ve Stapylococcus epidemidis İCA1 gen ifadesi üzerine etkilerinin karşilaştirilmasi

This study describes a comparison of latex-silicone coated, polyvinyl chloride, polyvinylpyrolidone coated polyurethane and polyurethane catheters for the delivery of adhesion genes in an in vitro biofilm model. Aglutinin-like sequence (ALS1), hypha-specific surface protein (HWP1) genes of Candida albicans and intracellular adhesion (ICA1) gene for Staphylococcus epidermidis were targeted. Four different biofilm models were constructed on four different bio-materials. Biofilm mass was separeted from the surface of the catheter and genomic mRNA was isolated. It was quantified in a real time PCR assay. ALS1 and HWP1 gene copy numbers were found to be 2.50x105, 1.30x104 copy/ml, respectively, and ICA1 gene copy number was found to be 1.80x105 copy/ml on the biofilm of the urinary catheters latex-silicone coated material. ALS1 and HWP1 gene copy numbers were found to be 7.56x106, 6.88x105 copy/ml, respectively, and ICA gene copy number was found to be 4.20x105 copy/ml on the biofilm removed from endotracheal tube, made from polyvinyl chloride. biofilm formation was not seen and biofilm associated C.albicans ALS1, HWP1 and S.epidermidis ICA genes were not expressed on the surface of nasogastric tube made from polyvinylpyrolidone coated polyurethane and the aspiration catheter made from polyurethane. In the PVP slime model, mean copy number was 2.70x102, 2.27x101 and 2.30x101 for ALS1, HWP1 and ICA, respectively. Mean copy number of adhesion genes on PUC were 1.50x101, 1.20x101 and 3.10x101 for ALS1, HWP1 and ICA, respectively. Adhesion genes were found as expressed in higher levels on the polyvinyl chloride catheter than those in siliconized one. Minimum gene expression levels were detected in polyvinylpyrolidone coated polyurethane and polyurethane catheters. It may result in the preferation of one material from other in the clinical use because of the different gene expression levels on the surface of these biomaterials.

PDF

___

1. Arciola CR, Campoccia D, Gamberini S, Cervellati M, Donati E, Montanaro L: Detection of slime production by means of an optimised Congo red agar plate test based on a colourimetric scale in Staphylococcus epidermidis clinical isolates genotyped for ica locus, Biomaterials 2002;23(21): 4233-9.
2. Cafiso V, Bertuccio T, Santagati M et al: Presence of the ica operon in clinical isolates of Staphylococcus epidermidis and its role in biofilm production, Clin Microbiol Infect 2004;10(12):1081-8.
3. Cheng G, Wozniak K, Wallig MA, Jr. Fidel PJ, Trupin SR, Hoyer LL: Comparison between Candida albicans agglutinin-like sequence gene expression patterns in human clinical specimens and models of vaginal candidiasis, Infect Immun 2005;73(3):1656-63.
4. Christensen GD, Simpson WA, Bisno AL, Beachey EH: Adherence of slime-producing strains of Staphylococcus epidermidis to smooth surfaces, Infect Immun 1982;37(1):1318-26.
5. Francois P, Vaudaux P, Nurdin N, Mathieu HJ, Descouts P, Lew DP: Physical and biological effects of a surface coating procedure on polyurethane catheters, Biomaterials 1996;17(7):667-78.
6. Ghannoum MA, O’Toole GA (Eds): Microbial Biofilms, ASM Press, Washington, DC (2004).
7. Hancock V, Klemm P: Global gene expression profiling of asymptomatic bacteriuria Escherichia coli during biofilm growth in human urine, Infect Immun 2007;75(2):966-76.
8. Henrickson BKJ, Axtell RA, Hoover SM et al: Prevention of central venous catheter-related infections and thrombotic events in immunocompromised children by the use of vancomycin/ ciprofloxacin/heparin flush solution: A randomized, multicenter, double-blind trial, J Clin Oncol 2000;18(6):1269-78.
9. Hoyer LL, Green CB, Oh SW, Zhao X: Discovering the secrets of the Candida albicans agglutinin-like sequence (ALS) gene family - a sticky pursuit, Med Mycol 2008;46(1):1-15.
10. Kumamoto CA, Vinces MD: Contributions of hyphae and hypha-co-regulated genes to Candida albicans virulence, Cell Microbiol 2005;7(11):1546-54.
11. Li H, Xu L, Wang J, Wen Y, Vuong C, Otto M, Gao Q: Conversion of Staphylococcus epidermidis strains from commensal to invasive by expression of the ica locus encoding production of biofilm exopolysaccharide, Infect Immun 2005;73(5):3188-91.
12. Lopez-Lopez G, Pascual A, Perea EJ: Effect of plastic catheter material on bacterial adherence and viability, J Med Microbiol 1991;34(6):349-53.
13. Mehall JR, Saltzman DA, Jackson RJ, Smith SD: Catheter materials affect the incidence of late blood-borne catheter infection, Surg Infect (Larchmt) 2001;2(3):225-30.
14. Ninin E, Caroff N, Espaze E et al: Assessment of ica operon carriage and biofilm production in Staphylococcus epidermidis isolates causing bacteraemia in bone marrow transplant recipients, Clin Microbiol Infect 2006;12(5):446-52.
15. Nobile CJ, Mitchell AP: Genetics and genomics of Candida albicans biofilm formation, Cellular Microbiol 2006;8(9):1382-91.
16. O’Gara JP: ica and beyond: biofilm mechanisms and regulation in Staphylococcus epidermidis and Staphylococcus aureus, FEMS Microbiol Lett 2007;270(2):179-88.
17.Ruggeri V, Francolini I, Donelli G, Piozzi A; Synthesis, characterization, and in vitro activity of antibiotic releasing polyurethanes to prevent bacterial resistance, j Biomed Mater Res A 2007;81(2):287-98.
18. Warren DK, Quadir WW, Hollenbeak CS, Edward AM, Cox MJ, Frase VJ: Attributable cost of catheter-associated bloodstream infections among intensive care patients in a nonteaching hospital, Grit Care Med 2006;34(8):2243-4.
19. Yeater KM, Chandra J, Cheng G et al: Temporal analysis of Candida albicans gene expression during biofilm development, Microbiology 2007;153(8):2373-85.