Experimental Grounds for Using Collagen-Based Anti-Adhesion Barrier Coated with Biocides for Prevention of Abdominal Surgical Infection

The aim of the study was to evaluate adhesive properties of the surgical anti-adhesion barrier based on collagen in combination with 0.05% Chlorhexidine bigluconate solution and Prontosan in experiment performed in vitro.

Materials and Methods. The study was carried out using CollaGUARD adhesion barrier consisting of renatured horse type I collagen and reference strains of Еscherichia coli АТСС^®25922, Klebsiella pneumoniae АТСС^®700603, Pseudomonas aeruginosa АТСС^®27853, Staphylococcus aureus АТСС^®25923. Bacteria adhesion to the barrier (membrane) untreated and combined with biocides was evaluated by determining the viable colony forming unit (CFU/cm²) numbers and using atomic force microscopy after 24 h and 6 days.

Results. Cells of all bacterial strains adhered to the surface of the membrane within 24 h forming a biofilm of bacilli tightly adjacent to each other (E. coli, P. aeruginosa) or cocci (S. aureus) integrated between disorganized collagen fibers, or adherent bacteria were seen separately (K. pneumoniae). The Sq index characterizing the surface roughness of the biofilm formed by S. aureus bacteria was 221.3±38.6 nm and was 3.0, 3.8, and 3.6 times higher compared with that for E. coli (72.8±12.6 nm), K. pneumoniae (57.5±21.8 nm), and P. aeruginosa (60.8±22.1 nm), respectively, and also exceeded 4.4 times the Sq index for the membrane itself (50.3±26.3 nm). Collagen degradation was revealed in case of contamination by protease-producing bacteria with collagenase activity: P. aeruginosa for 24 h and S. aureus for 6 days. Adsorption of the biocide on the membrane surface after its short exposure to Chlorhexidine bigluconate solution or Prontosan led to inhibited growth and adhesion of cells of bacteria, except for P. aeruginosa. In experimental models, in agar medium and in suspension culture, Chlorhexidine bigluconate proved to be more effective than Prontosan. The difference between the biocides in inhibiting the adhesion of bacteria to the membrane surface is not associated with changes in its surface roughness.

Conclusion. Impregnation of a surgical membrane with antibacterial compounds just before the implantation can serve as an additional method for preventing abdominal surgical infection.

1. Kondratovich L. The basics for comprehension of adhesive process formation in abdominal cavity. Perioperative prevention by means of anti-adhesive drugs (review of literature). Journal of New Medical Technologies 2014; 21(3): 169–173, https://doi.org/10.12737/5929.
2. Nazarenko A.A., Akimov V.P. Overcoming of adhesiogenesis. Medical Herald of the South of Russia 2014; 4: 15–18.
3. Ayushinova N.I., Shurygina I.A., Shurygin M.G., Panasuk A.I. Contemporary approaches to the prevention of intraperitoneal adhesions. Sibirskiy meditsinskiy zhurnal 2011; 105(6): 16–20.
4. Snegirev I.I., Mironov V.I., Bashlykov D.V. Nontumoral acute ileus: diagnosis and treatment. Sibirskiy meditsinskiy zhurnal 2010; 99(8): 163–165.
5. Bilsel Y., Abci I. The search for ideal hernia repair; mesh materials and types. Int J Surg 2012; 10(6): 317–321, https://doi.org/10.1016/j.ijsu.2012.05.002.
6. Brochhausen C., Schmitt V.H., Planck C.N., Rajab T.K., Hollemann D., Tapprich C., Krämer B., Wallwiener C., Hierlemann H., Zehbe R., Planck H., Kirkpatrick C.J. Current strategies and future perspectives for intraperitoneal adhesion prevention. J Gastrointest Surg 2012; 16(6): 1256–1274, https://doi.org/10.1007/s11605-011-1819-9.
7. Koc O., Duran B., Topcuoglu A., Bugdayci G., Yilmaz F., Dönmez M. Intraperitoneal administration of single dose type I collagen or low dose melatonin to prevent intraperitoneal adhesion formation: а comparative study. Eur J Obstet Gynecol Reprod Biol 2009; 145(2): 209–213, https://doi.org/10.1016/j.ejogrb.2009.05.019.
8. Sullivan E.K., Kamstock D.A., Turner A.S., Goldman S.M., Kronengold R.T. Evaluation of a flexible collagen surgical patch for reinforcement of a fascial defect: experimental study in a sheep model. J Biomed Mater Res B Appl Biomater 2008; 87(1): 88–94, https://doi.org/10.1002/jbm.b.31073.
9. Lee C.H., Singla A., Lee Y. Biomedical applications of collagen. Int J Pharm 2001; 221(1–2): 1–22, h ttps://doi.org/10.1016/s0378-5173(01)00691-3.
10. Gorskij V.A., Sivkov A.S., Agapov M.A., Titkov B.E., Schadskij S.O. The first experience of using a single-layer intra-abdominal collagen plate. Khirurgiia. Zhurnal im. N.I. Pirogova 2015; 5: 59–61, https://doi.org/10.17116/hirurgia2015559-61.
11. Schönleben F., Reck T., Tannapfel A., Hohenberger W., Schneider I. Collagen foil (TissuFoil E) reduces the formation of adhesions when using polypropylene mesh for the repair of experimental abdominal wall defects. Int J Colorectal Dis 2006; 21(8): 840–846, https://doi.org/10.1007/s00384-006-0091-z.
12. Instruction manual for medical device “KolGARA absorbable adhesion prevention collagen barrier (membrane)”. 2013.
13. Pérez-Köhler B., García-Moreno F., Bayon Y., Pascual G., Bellón J.M. Inhibition of Staphylococcus aureus adhesion to the surface of a reticular heavyweight polypropylene mesh soaked in a combination of chlorhexidine and allicin: an in vitro study. PLoS One 2015; 10(5): e0126711, https://doi.org/10.1371/journal.pone.0126711.
14. Liang H.-C., Chang Y., Hsu C.-K., Lee M.-H., Sung H.-W. Effects of crosslinking degree of an acellular biological tissue on its tissue regeneration pattern. Biomaterials 2004; 25(17): 3541–3552, https://doi.org/10.1016/j.biomaterials.2003.09.109.
15. Siluyanov S.V., Aliev S.R. First experience in the use of adhesion prevention collagen membrane during abdominal and pelvic surgery. Russkij medicinskij zurnal 2015; 23(13): 789–795.
16. Birkenhauer E., Neethirajan S., Weese J.S. Collagen and hyaluronan at wound sites influence early polymicrobial biofilm adhesive events. BMC Microbiology 2014; 14(1): 191, https://doi.org/10.1186/1471-2180-14-191.
17. García-Pumarino R., Pascual G., Rodríguez M., Pérez-Köhler B., Bellón J.M. Do collagen meshes offer any benefits over preclude® ePTFE implants in contaminated surgical fields? A comparative in vitro and in vivo study. J Biomed Mater Res B Appl Biomater 2014; 102(2): 366–375, https://doi.org/10.1002/jbm.b.33015.
18. Trafny E.A., Kowalska K., Grzybowski J. Adhesion of Pseudomonas aeruginosa to collagen biomaterials: effect of amikacin and ciprofloxacin on the colonization and survival of the adherent organisms. J Biomed Mater Res 1998; 41(4): 593–599, https://doi.org/10.1002/(sici)1097-4636(19980915)41:4593::aid-jbm113.0.co;2-g.
19. Clarke S.R., Foster S.J. Surface adhesins of Staphylococcus aureus. Adv Microb Physiol 2006; 51: 187–224, https://doi.org/10.1016/s0065-2911(06)51004-5.
20. Plotkowski M.C., Chevillard M., Pierrot D., Altemayer D., Zahm J.M., Colliot G., Puchelle E. Differential adhesion of Pseudomonas aeruginosa to human respiratory epithelial cells in primary culture. J Clin Invest 1991; 87(6); 2018–2028, https://doi.org/10.1172/jci115231.
21. Stepińska M., Trafny E.A. Modulation of Pseudomonas aeruginosa adherence to collagen type I and type II by carbohydrates. FEMS Immunol Med Microbiol 1995; 12(3–4): 187–194, https://doi.org/10.1016/0928-8244(95)00066-6.
22. Pouttu R., Puustinen T., Virkola R., Hacker J., Klemm P., Korhonen T.K. Amino acid residue Ala-62 in the FimH fimbrial adhesion is critical for the adhesiveness of meningitis-associated Escherichia coli to collagens. Mol Microbiol 1999; 31(6): 1747–1757, https://doi.org/10.1046/j.1365-2958.1999.01311.x.
23. Rêgo A.T., Johnson J.G., Gelbel S., Enguita F.J., Clegg S., Waksman G. Crystal structure of the MrkD1P receptor binding domain of Klebsiella pneumoniae and identification of the human collagen V binding interface. Mol Microbiol 2012; 86(4): 882–893, https://doi.org/10.1111/mmi.12023.
24. Sadava E.E., Krpata D.M., Gao Y., Novitsky Y.W., Rosen M.J. Does presoaking synthetic mesh in antibiotic solution reduce mesh infections? An experimental study. J Gastrointest Surg 2013; 17(3): 562–568, https://doi.org/10.1007/s11605-012-2099-8.
25. Wiegering A., Sinha B., Spor L., Klinge U., Steger U., Germer C.T., Dietz U.A. Gentamicin for prevention of intraoperative mesh contamination: demonstration of high bactericide effect (in vitro) and low systemic bioavailability (in vivo). Hernia 2014; 18(5): 691–700, https://doi.org/10.1007/s10029-014-1293-x.
26. Yabanoğlu H., Arer İ.M, Çalıskan K. The effect of the use of synthetic mesh soaked in antibiotic solution on the rate of graft infection in ventral hernias: a prospective randomized study. Int Surg 2015; 100(6): 1040–1047, https://doi.org/10.9738/intsurg-d-14-00304.1.
27. Abdominal’naya khirurgicheskaya infektsiya (klassifikatsiya, diagnostika, antimikrobnaya terapiya). Rossiyskie natsional’nye rekomendatsii [Abdominal surgical infection. Russian national recommendations]. Pod red. Savel’eva V.S., Gel’fanda B.R. [Savel’ev V.S., Gel’fand B.R. (editors)]. Moscow: OOO “Kompaniya Borges”; 2011.
28. Morrissey I., Hackel M., Badal R., Bouchillon S., Hawser S., Biedenbach D. A review of ten years of the study for monitoring antimicrobial resistance trends (SMART) from 2002 to 2011. Pharmaceuticals 2013; 6(11): 1335–1346, https://doi.org/10.3390/ph6111335.
29. Sartelli M., Catena F., Ansaloni L., Coccolini F., Corbella D., Moore E.E., Malangoni M., Velmahos G., Coimbra R., Koike K., Leppaniemi A., Biffl W., Balogh Z., Bendinelli C., Gupta S., Kluger Y., Agresta F., Di Saverio S., Tugnoli G., Jovine E., Ordonez C.A., Whelan J.F., Fraga G.P., Gomes C.A., Pereira G.A., Yuan K.C., Bala M., Peev M.P., Ben-Ishay O., Cui Y., Marwah S., Zachariah S., Wani I., Rangarajan M., Sakakushev B., Kong V., Ahmed A., Abbas A., Gonsaga R.A., Guercioni G., Vettoretto N., Poiasina E., Díaz-Nieto R., Massalou D., Skrovina M., Gerych I., Augustin G., Kenig J., Khokha V., Tranà C., Kok K.Y., Mefire A.C., Lee J.G., Hong S.K., Lohse H.A., Ghnnam W., Verni A., Lohsiriwat V., Siribumrungwong B., El Zalabany T., Tavares A., Baiocchi G., Das K., Jarry J., Zida M., Sato N., Murata K., Shoko T., Irahara T., Hamedelneel A.O., Naidoo N., Adesunkanmi A.R., Kobe Y., Ishii W., Oka K., Izawa Y., Hamid H., Khan I., Attri A., Sharma R., Sanjuan J., Badiel M., Barnabé R. Complicated intra-abdominal infections worldwide: the definitive data of the CIAOW Study. World J Emerg Surg 2014; 9(1): 37, https://doi.org/10.1186/1749-7922-9-37.
30. Sanders D.L., Kingsnorth A.N., Lambie J., Bond P., Moate R., Steer J.A. An experimental study exploring the relationship between the size of bacterial inoculum and bacterial adherence to prosthetic mesh. Surg Endosc 2013; 27(3): 978–985, https://doi.org/10.1007/s00464-012-2545-4.
31. Caballero A., Thibodeaux B., Marquart M., Traidej M., O’Callaghan R. Pseudomonas keratitis: protease IV gene conservation, distribution, and production relative to virulence and other Pseudomonas proteases. Invest Ophthalmol Vis Sci 2004; 45(2): 522–530, https://doi.org/10.1167/iovs.03-1050.
32. Kida Y. Roles of Pseudomonas aeruginosa-derived proteases as a virulence factor. Nihon Saikingaku Zasshi 2013; 68(4): 313–323, https://doi.org/10.3412/jsb.68.313.
33. Leidal K.G., Munson K.L., Johnson M.C., Denning G.M. Metalloproteases from Pseudomonas aeruginosa degrade human RANTES, MCP-1, and ENA-78. J Interferon Cytokine Res 2003; 23(6): 307–318, https://doi.org/10.1089/107999003766628151.
34. Bailey A.J. The fate of collagen implants in tissue defects. Wound Repair Regen 2000; 8(1): 5–12, https://doi.org/10.1046/j.1524-475x.2000.00005.x.
35. Ohbayashi T., Irie A., Murakami Y., Nowak M., Potempa J., Nishimura Y., Shinohara M., Imamura T. Degradation of fibrinogen and collagen by staphopains, cysteine proteases released from Staphylococcus aureus. Microbiology 2011; 157(3): 786–792, https://doi.org/10.1099/mic.0.044503-0.
36. Suphatharaprateep W., Cheirsilp B., Jongjareonrak A. Production and properties of two collagenases from bacteria and their application for collagen extraction. N Biotechnol 2011; 28(6): 649–655, https://doi.org/10.1016/j.nbt.2011.04.003.
37. Zhang Y.Z., Ran L.Y., Li C.Y., Chen X.L. Diversity, structures, and collagen-degrading mechanisms of bacterial collagenolytic proteases. Appl Environ Microbiol 2015; 81(18): 6098–6107, https://doi.org/10.1128/aem.00883-15.
38. Kuznetsova M.V., Encheva Yu.A., Samartsev V.A. Influence of chlorhexidine and prontosan on dual species and monospecies biofilms formed by Staphylococcus aureus and Pseudomonas aeruginosa. Antibiotiki i khimioterapiya 2015; 60(11–12): 15–22.
39. Pirog T.P., Konon A.D., Beregovaya K.A., Shulyakova M.A. Antiadhesive properties of the surfactants of Acinetobacter calcoaceticus IMB B-7241, Rhodococcus erythropolis IMB Ac-5017, and Nocardia vaccinii IMB B-7405. Microbiology 2014; 83(6): 732–739, https://doi.org/10.1134/s0026261714060150.
40. Parshikov V.V., Samsonov A.A., Romanov R.V., Samsonov A.V., Gradusov V.P., Treushnikov V.V., Uspenskiy I.V., Sorokina O.V. Operative treatment of strangulated hernias with a use of reticular endoprostheses. Nizegorodskij medicinskij zurnal 2008; 6: 19–22.
41. Gristina A.G., Naylor P., Myrvik Q. Infections from biomaterials and implants: a race for the surface. Med Prog Technol 1988; 14(3–4): 205–224.

Kuznetsova M.V., Kuznetsova M.P., Afanasyevskaya E.V., Samartsev V.A. Experimental Grounds for Using Collagen-Based Anti-Adhesion Barrier Coated with Biocides for Prevention of Abdominal Surgical Infection. Sovremennye tehnologii v medicine 2018; 10(2): 66, https://doi.org/10.17691/stm2018.10.2.07