Frequency of bap and cpaA virulence genes in drug resistant clinical isolates of Acinetobacter baumannii and their role in biofilm formation

Document Type: Original Article

Authors

1 Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran

2 Student Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran

3 Infectious and Tropical Diseases Research Center, Tabriz University of Medical Sciences, Tabriz, Iran

4 Department of Microbiology, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran

5 Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran

Abstract

Objective(s): Acinetobacter baumannii has a high propensity to form biofilm and frequently causes medical device-related infections with multiple-drug-resistance in hospitals. The aim of this work is to study antimicrobial resistance and the role of bap and cpaA genes in biofilm formation by A. baumannii to understand how this pathogen persists in the hospital environment.
Materials and Methods: Theantibiotic resistance profile and in vitro biofilm-forming ability of one hundred clinical isolates of A. baumannii was evaluated by disc diffusion and crystal-violet staining methods, respectively. Isolates were tested for the presence of bap and cpaA genes.
Results: The isolates were highly resistant to cefepime, third-generation cephalosporins, ciprofloxacin, cotrimoxazole, aminoglycosides and carbapenems. Moreover, four isolates were resistant to colistin. Quantification of biofilm showed that 43% of the isolates were strong biofilm-producer. Furthermore, 32% of the isolates exhibited moderate biofilm-formation and showed initial binding activity. Frequency of bap and cpaA were determined 92% and 36%, respectively.
Conclusion: There was strong association between the presence of bap gene and biofilm formation by A. baumannii isolates (P=0.003). In addition, multidrug resistant isolates produced stronger biofilm than other isolates (P=0.0001). These results indicate importance of biofilm in resistance of isolates and effect of presence of bap gene in biofilm formation by A. baumannii strains.

Keywords


1.Durante-Mangoni E, Utili R, Zarrilli R. Combination therapy in severe Acinetobacter baumannii infections: an update on the evidence to date. Future Microbiol 2014; 9:773-789.

2.Falagas ME, Bliziotis IA, Siempos II. Attributable mortality of Acinetobacter baumannii infections in critically ill patients: a systematic review of matched cohort and case-control studies. Crit Care 2006; 10:R48.

3.Hall-Stoodley L, Costerton JW, Stoodley P. Bacterial biofilms: from the natural environment to infectious diseases. Nat Rev Microbiol 2004; 2:95-108.

4.Singhai M, Malik A, Shahid M, Malik MA, Goyal R. A study on device-related infections with special reference to biofilm production and antibiotic resistance. J Glob Infect Dis 2012; 4:193.

5.Sutherland IW. The biofilm matrix–an immobilized but dynamic microbial environment. Trends Microbiol 2001; 9:222-227.

6.Stoodley P, Sauer K, Davies D, Costerton JW. Biofilms as complex differentiated communities. Ann Rev Microbiol 2002; 56:187-209.

7.Patel SJ, Oliveira AP, Zhou JJ, Alba L, Furuya EY, Weisenberg SA, et al. Risk factors and outcomes of infections caused by extremely drug-resistant gram-negative bacilli in patients hospitalized in intensive care units. Am J Infect Control 2014; 42:626-631.

8.Luo TL, Rickard AH, Srinivasan U, Kaye KS, Foxman B. Association of blaOXA-23 and bap with the persistence of Acinetobacter baumannii within a major healthcare system. Front Microbiol 2015; 6:182.

9.Lasa I, Penadés JR. Bap: a family of surface proteins involved in biofilm formation. Res Microbiol 2006; 157:99-107.

10.Latasa C, Solano C, Penadés JR, Lasa I. Biofilm-associated proteins. C R Biol 2000; 329:849-857.

11.Brossard KA, Campagnari AA. The Acinetobacter baumannii biofilm-associated protein plays a role in adherence to human epithelial cells. Infect Immun 2012; 80:228-233.

12.Loehfelm TW, Luke NR, Campagnari AA. Identification and characterization of an Acinetobacter baumannii biofilm-associated protein. J Bacteriol 2008; 190:1036-1044.

13.Leung WS, Chu CM, Tsang KY, Lo FH, Lo KF, Ho PL. Fulminant community-acquired Acinetobacter baumannii pneumonia as a distinct clinical syndrome. Chest 2006; 129:102-109.

14.Soares AJ, Santos M, Trugilho M, Neves-Ferreira A, Perales J, Domont G. Differential proteomics of the plasma of individuals with sepsis caused by Acinetobacter baumannii. J Proteomics 2009; 73:267-278.

15.Delvaeye M, Conway EM. Coagulation and innate immune responses: can we view them separately? Blood 2009; 114:2367-2374.

16.Massberg S, Grahl L, von Bruehl M-L, Manukyan D, Pfeiler S, Goosmann C, et al. Reciprocal coupling of coagulation and innate immunity via neutrophil serine proteases. Nat Med 2010; 16:887-896.

17.Tilley D, Law R, Warren S, Samis JA, Kumar A. CpaA a novel protease from Acinetobacter baumannii clinical isolates deregulates blood coagulation. FEMS Microbiol Lett 2014; 356:53-61.

18.MacFaddin JF. Biochemical tests for identification of medical bacteria, 3rd ed. Philadelphia, PA: Lippincott Williams & Wilkins; 2000.

19.Woodford N, Ellington MJ, Coelho JM, Turton JF, Ward ME, Brown S, et al. Multiplex PCR for genes encoding prevalent OXA carbapenemases in Acinetobacter spp. Int J Antimicrob Agents 2006; 27:351-353.

20.Pajand O, Ahangarzadeh Rezaee M, Nahaei MR, Mahdian R, Aghazadeh M, Soroush MH, et al. Study of the carbapenem resistance mechanisms in clinical isolates of Acinetobacter baumannii: Comparison of burn and non-burn strains. Burns 2013; 39:1414-1419.

21.Clinical Laboratory Standards Institute, M100-S24: Performance Standards for Antimicrobial Susceptibility testing: Twenty-First Informational Supplement. Clinical Laboratory Standards Institute; Wayne, Pa, USA: 2014.

22.Ahangarzadeh Rezaee M, Abdinia B. Etiology and antimicrobial susceptibility pattern of pathogenic bacteria in children subjected to UTI: A referral hospital-based study in Northwest of Iran. Medicine 2015; 94:e1606.

23.Kafil HS, Mobarez AM, Moghadam M, Hashemi Z, MY. Gentamicin induces efaA expression and biofilm formation in Enterococcus faecalis. Microb Pathog 2016; 92:30-35.

24.Kafil HS, Mobarez AM. Assessment of biofilm formation by enterococci isolates from urinary tract infections with different virulence profiles. J King Saud Univ-Sci 2015; 27:312-7.

25.Akers KS, Mende K, Cheatle KA, Zera WC, Yu X, Beckius ML, et al. Biofilms and persistent wound infections in United States military trauma patients: a case-control analysis. BMC Infect Dis 2014; 14:190.

26.Peleg AY, Seifert H, Paterson DL. Acinetobacter baumannii: emergence of a successful pathogen. Clin Microbiol Rev 2008; 21:538-582.

27.Bergogne-Berezin E, Towner K. Acinetobacter spp. as nosocomial pathogens: microbiological, clinical, and epidemiological features. Clin Microbiol Rev 1996; 9:148-165.

28.Bialvaei AZ, Samadi Kafil H. Colistin, mechanisms and prevalence of resistance. Curr Med Res Opin 2015; 31:707-721.

29.Kollef MH, Fraser VJ. Antibiotic resistance in the intensive care unit. Ann Intern Med 2001; 134:298-314.

30.Soroush S, Haghi-Ashtiani MT, Taheri-Kalani M, Emaneini M, Aligholi M, Sadeghifard N, et al. Antimicrobial resistance of nosocomial strain of Acinetobacter baumannii in Children’s Medical Center of Tehran: a 6-year prospective study. Acta Med Iran 2010; 48:178-184.

31.Sohrabi N, Farajnia S, Akhi MT, Nahaei MR, Naghili B, Peymani A, et al. Prevalence of OXA-type β-lactamases among Acinetobacter baumannii isolates from Northwest of Iran. Microb Drug Resist 2012; 18:385-389.

32.Ghasemian R, Ahanjan M, Fatehi E, Shokri M. Prevalence and antibiotic resistance pattern of Acinetobacter isolated from patients admitted in ICUs in Mazandaran, Northern Iran. Glob J Health Sci 2016; 8:112-119.

33.Mohajeri P, Farahani A, Feizabadi MM, Ketabi H, Abiri R, Najafi F. Antimicrobial susceptibility profiling and genomic diversity of Acinetobacter baumannii isolates: A study in western Iran. Iran J Microbiol 2013; 5:195-202.

34.Azizi O, Shakibaie MR, Modarresi F, Shahcheraghi F. Molecular detection of class-D OXA carbapenemase genes in biofilm and non-biofilm forming clinical isolates of Acinetobacter baumannii. Jundishapur J Microbiol 2015; 8:e21042.

35.Lee HW, Koh Y, Kim J, Lee JC, Lee YC, Seol SY, et al. Capacity of multidrug-resistant clinical isolates of Acinetobacter baumannii to form biofilm and adhere to epithelial cell surfaces. Clin Microbiol Infect 2008; 14:49-54.

36.Rodriguez-Bano J, Marti S, Soto S, Fernández-Cuenca F,  Cisneros JM, Pachon J, et al. Biofilm formation in Acinetobacter baumannii: associated features and clinical implications. Clin Microbiol Infect 2008; 14:276-278.

37.Sánchez CJ, Mende K, Beckius ML, Akers KS, Romano DR, Wenke JC, et al. Biofilm formation by clinical isolates and the implications in chronic infections. BMC Infect Dis 2013; 13:47.

38.Cucarella C, Solano C, Valle J, Amorena B, Lasa Í, Penadés JR. Bap, a Staphylococcus aureus surface protein involved in biofilm formation. J Bacteriol 2001; 183:2888-2896.

39.Lasa I. Towards the identification of the common features of bacterial biofilm development. Intl Microbiol 2006; 9:21-28.

40.Badmasti F, Siadat SD, Bouzari S, Ajdary S, Shahcheraghi F. Molecular detection of genes related to biofilm formation in multidrug-resistant Acinetobacter baumannii isolated from clinical settings. J Med Microbiol 2015; 64:538-543.

41.Goh HS, Beatson SA, Totsika M, Moriel DG, Phan M-D, Szubert J, et al. Molecular analysis of the Acinetobacter baumannii biofilm-associated protein. Appl Environ Microbiol 2013; 79:6535-6543.