Original article / research
Year :
2013 |
Month :
April
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Volume :
2 |
Issue :
1 |
Page :
12 - 15 |
Full Version
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Isolation Speciation and Determination of High Level Aminoglycoside Resistance
of Enterococci Among Hospitalised Patients in Davangere
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K. Suresh, B. Saipriya, G. Viswanath 1. MD Professor
2. MBBS Post graduate
3. MD, Professor & HOD, Department of Microbiology, J.J.M. Medical College, Davangere, Karnataka, India.
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Correspondence
Address :
K. Suresh, B. Saipriya, G. Viswanath, Dr. K. Suresh,
Professor, Department of Microbiology,
JJM Medical College, Davangere,
Karnataka-577004, India.
Ph: 09986048719
Email: dr.sureshk@rediffmail.com
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| ABSTRACT |  | : Background: Enterococcus, considered a normal commensal of intestinal tract is one of the fast emerging pathogen causing serious and life threatening hospital borne infections. This is attributed to acquisition of multi-drug resistance.
Aims and Objectives: The present study was undertaken to isolate and characterize Enterococci from clinical specimens and determine the anti-microbial susceptibility pattern of these isolates.
Methods: A total of 58 Enterococcal isolates from various clinical samples were speciated as per the scheme of Facklam and Collins. Anti-bacterial susceptibility pattern was determined by Kirby Bauer disc diffusion method with recommended drugs including high level aminoglycoside resistance. MIC for Vancomycin and Gentamicin was determined by E – test method.
Results: Of the total 58 Enterococcal isolates, 52 were E. faecalis and 6 E. faecium. Antibiotic susceptibility tests showed high level resistance to penicillin, Ampicillin, Gentamicin and Streptomycin. All strains were sensitive to Vancomycin, Linezolid, Teicoplanin, E. faecium was more resistant than E. faecalis to the tested antibiotics. MIC for vancomycin was in range 1-4μg/ml.
Conclusions: We conclude that Enterococcal strains with high rate of resistance to penicillin and aminoglycosides are prevalent in our nosocomial setting. Therefore, there is an urgent need for more rational and restricted use of antimicrobials in order to minimize the selection and spread of such strains. |
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Keywords
: Enterococci, High level aminoglycoside resistance (HLAR), MIC |
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DOI and Others
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INTRODUCTION |
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Enterococci, an indigenous flora of the intestinal tract, oral cavity and the genito urinary tract of the humans and animals, are known to be relatively avirulent in healthy individuals, but have become important opportunistic pathogens, especially in hospitalized patients (1). Recent years have witnessed increased interest in Enterococci not only because of their ability to cause serious infections like endocarditis, bacteremia, intra-abdominal and urinary tract infection, but also because of their increasing resistance to many anti-microbial agents (2). Infections by Enterococci have traditionally been treated with cell wall active agents in combination with an aminoglycoside, however emergence ofhigh level resistance to aminoglycosides, β lactam antibiotics and to vancomycin by some strains has led to the failure of synergistic effects of combination therapy (3), (4).
Although 23 species in genus Enterococcus have been recognized, most common species are E. faecalis followed by E. faecium. E. faecium is more resistant species than E. faecalis and emergence of vancomycin resistance in it has caused an increase in frequency of its isolations (5). Nevertheless, the incidence of other species of Enterococci is underestimated because of frequent misidentification (6). Hence proper identification to species level is essential for proper management and prevention of this bacterial infection in any health care institution.
Hence the present study was conducted to know the species prevalence and the high level aminoglycoside resistance of Enterococcal isolates.
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Material and Methods |
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The present study was conducted in the Department of Microbiology from June 2011 – June 2012. Various clinical specimens such as urine, pus, abdominal drain fluid and blood were processed for the isolation of Enterococci. The specimens were also plated on blood agar and Mac Conkey’s agar for the isolation of concomitant organisms along with Enterococci. Enterococci were identified on the basis of characteristic colony morphology, gram staining and catalase test, and confirmed by Bile – esculin hydrolysis, salt – tolerance, heat tolerance tests. Enterococcal strains were further identified to the species level by using the conventional physiological tests devised by Facklam and Collins (7). Anti-microbial susceptibility testing was done by Kirby – Bauer disc diffusion method as per the recommendations of CLSI (8). The MIC of vancomycin was determined by E-test. The source of media and antibiotic discs were Hi-media ltd. Standard strains E. faecalis ATCC 29212 was used as control.
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Results |
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Result of the 250 samples tested, comprising urine – 120, pus – 75, body fluids – 30 and blood – 25, a total of 58 enterococcal isolates 52 –E. faecalis (89.6%) and 6 – E. faecium (10.4%) were obtained (Table/Fig 1). 50 isolates were pure culture of Enterococci, remaining 8 isolates were associated with E.coli, Proteus, Pseudomonas and Staphylococcus aureus.The majority of isolates were from urine 36 (62%), E. faecalis 34, E. faecium 2, 6 from non surgical wounds (10.3%), 8 from surgical wound (13.8%), 6 from blood (10.3%) and 2 from body fluids (3.4%). Both E. faecalis and E. faecium exhibited 100% sensitivity for Vancomycin, Linezolid, Teicoplanin. Nitrofurantoin was used only for urinary isolates and was 100% sensitive. The resistence pattern of E. faecalis: Ampicillin (54%), Penicillin (35%), Ciprofloxacin (54%), Ofloxacilin (54%), Gentamicin (54%), Streptomycin (31%) (Table/Fig 2).
High level gentamicin disc (120μg) and high level streptomycin disc (300μg) were used for detection of high level aminoglycoside resistance in Enterococci. Among the 58 isolates, 28 strains of E. faecalis exhibited HLGR (53.8%) and 8 strains showed both HLGR and HLSR (15.3%) 20 strains showed only HLGR and was sensitive to streptomycin (38.4%). The remaining 24 isolates of E. faecalis were sensitive to gentamicin (46%). Among 6 isolates of E. faecium 4 strains exhibited HLGR (66.6%) (Table/Fig 3).
High level streptomycin disc (300μg) detected totally 16 HLSR strains (30.76%), 8 strains were both HLGR and HLSR, 8 showed only HLSR (15.3%) among E. faecalis isolates. (Table/Fig 3) 2 isolates of E. faecium (33.3%) exhibited HLSR. MIC of Gentamicin was performed by E-test. The strip with MIC range (0.064-1024μg) was used. 28 isolates of E. faecalis and 4 isolates of E. faecium had MIC > 512μg (resistant). MIC of vancomycin determined by E-test, strips with MIC range (0.016-256μg) was used. All 58 isolates showed MIC < 4μg (sensitive).
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Discussion |
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Enterococci are commensals of the gastrointestinal tract of human beings. Over the past 2 decades they have becomeimportant nosocomial pathogens probably due to inherent resistance to antibiotics (cephalosporins), ability to adhere to indwelling medical devices, and ability to survive adverse environmental conditions (9). Correct speciation is very important since there is variation in resistance to antibiotics by particular enterococcal species (10). In this study E. faecalis is the predominant species isolated (89.61%) followed by E.faecium (10.4%) in accordance with other studies of Gary Cotter et al., (11); Simonson et al., (12), Mohammad Rahbar (13). In contrary Baragundi Mahesh et al., (14) reported E. faecium as the predominant species (47.50%). Majority of the isolates were from urine, similar to other studies (15), (16).
This study showed E. faecium to be more resistant to ampicillin than E. faecalis. this correlates with the study by Jyotsna Agarwal et al., (17) and Steven Gordon et al., (18). Penicillin resistance was more in E.faecium than E.faecalis. In contrast L.A Sechi et al., reported 60% of E. faecalis isolates resistant to penicillin (19). This study showed 100% sensitivity to vancomycin, teicoplanin and Linezolid, similar to results observed in other studies (20),(16). High level gentamicin resistance was seen in both E. faecalis and E.faecium, consistent with other studies (21), (22). In our study among E. faecalis and E. faecium isolates resistant pattern was more for gentamicin than streptomycin, which is similar to previous studies (23), (24). E. faecium strains were observed to be more resistant to the tested antimicrobials in accordance to studies from India and outside (20), (21). Even though there are studies suggestive of arising resistance pattern to vancomycin in Enterococci (25), (3) no such pattern was observed in our isolates and isolates have MIC range between 1-4μg/ml for vancomycin.
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Conclusion |
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This study showed increasing high level aminoglycoside resistence in hospital setting.Therfore emphasising the rational and restricted use of antibiotics to minimise the selection and spread of such strains.
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| 1. | Murray BE. The life and times of Enterococci. Clin Microbiol Rev. 1990; 3: 46-65. | 2. | Moellering RC. Jr Emergence of Enterococci as a significant pathogen. Clin Infect Dis. 1992; 14: 1173-78. | 3. | Jesudason MV, Pratima VL, Pandian R,l Abigail S. Characterization of penicillin resistant enterococci. Indian J Med Microbiol. 1998; 16(1): 16-8. | 4. | Patterson JE, Zervos M. High level gentamicin resistance in Enterococcius, microbiology, genetic basis and epidemiology. Rev Infect Dis. 1990; 12: 644-51. | 5. | Rice LB. Emergence of Vancomycin resistant Enterococci EmergInfect Dis 2001; 7: 183-87. | 6. | Vittal P Prakash, Sambasiva R Rao, Subhash C Parija. Emergence of unusual species of enterococci causing infections, South India. BMC Infect disease. 2005 Mar; 5:14. | 7. | Facklam RR, Collins MD. Identification of enterococcus species isolated from human infection by a conventional test scheme. J Clin Microbiol. 1989; 27: 731-34. | 8. | Clinical and Laboratory standards Institute. Performance Standards for Antimicrobial susceptibility testing; 20th informational supplement. CLSI/NCCLS M100-S20. Wayne(PA). The Institute; 2010. | 9. | Shabnam Qamer, Jonathan Sandoe, Kevin G Kerr. Use of colony morphology to distinguish different Enterococcal strains and species in mixed culture from clinical specimens. Journ. of Clin Micro. June 2003: p 2644-46. | 10. | PJ Desai, D Pandit, M Mathur, A Gogate. Prevalence, identification and distribution of various species of enterococci isolated from clinical specimens with special reference to UTI in catheterized patients. Indian journal of Medical Microbiology. 2001; 19(3): 132-37. | 11. | Gary Cotter and Catherine C Adley. Comparison and evaluation of antimicrobial susceptibility testing of Enterococci performed in accordance with six National Committee standardized disk diffusion procedures. Journal of Clinic Micro. Oct 2001; p.3753-56. | 12. | G.S Simonson, L Smabrekkel, DL Monnet, TL Sorensen, JK Moller, et al. KG Kristensson et al. Prevalence of resistance to ampicillin, gentamicin and vancomycin in Enterococcus faecalis and E.facium isolates from clinical specimens and use of antimicrobials in five Nardic hospitals. Journal of Antimicrobial chemotherapy. (2003) 51, 323-31. | 13. | Mohammed Rahbar, Massoud Hajia, Mohammad Farzaneh Khan. Activity of Nitrofurantoin against isolates of vancomycin – resistant enterococci (VRE): A three layer survey in an Iranian hospital. Iran journal of patho. Nov. 2007; vol.2(4). | 14. | Baragundi Mahesh, Suresh B Sonth, Shivakumar S. Solabannavar, Chidanand S. Patil, Vishwanath, Yemul, Species prevalence and antimicrobial resistance pattern of enterocoocal isolates in a tertiary health care centre. Journal of clinical and diagnostic research. 2010 Dec; (4): 3405-09. | 15. | Maria Bitsori, Sofia Maraki, Maria Raissaki, Anna Bakantaki, Emmannouil Galanakis. Community acquired enterococcal urinary tract infections. Paed Nephro. 2005; 20: 1583-86. | 16. | Neelam Taneja, Phulan Rani, Rekha Emmanuel, Meera Sharma. Significance of vancomycin resistant enterococci from urinary specimens at a tertiary care centre in northern India. Indian J Med Res. 2004 Feb: pp. 72-74. | 17. | Jyotsna Agarwal, Rajkumar Kalyan and Mastan Singh. High level aminoglycoside resistance and β lactamase production in Enterococci at a tertiary care hospital in India. Jpn. J. Infect. Dis. 2009; 62: 158-59. | 18. | Steven Gordon, Jana M Swenson, Bertha C Hill, Nan E Pigott, RR Facklam. Anti-microbial susceptibility patterns of common and unusual species of Enterococci causing infections in the United States. Sep. 1992; vol. 30, no. 9: p. 2373-78. | 19. | I Dupre, S Zanetti, AM Schito, G Fadda, LA Sechi. Incidence of cirulence determinants in clinical enterococcus faecium and enterococcus faecalis isolates collected in Sardina, Journal Med Microbiol. 2003; 52: 491-98. | 20. | Srujana Mohanty, Swapna Jose, Ritu Singhal, Seema Sood, Benu Dhawan et al. Bimal K Das et al. species prevalence and antimicrobial susceptibility testing of clinical isolates of Enterococcus faecium and Enterococcus faecalis. Journ of clin. Microbiology. 1992; vol. 30: (1) p. 41-45. | 21. | M Louie, AE Simor, S Szeto, M Patel, B Kreiswirth, D E Low.Susceptibility testing of clinical isolates of enterococcus facium and enterococcus faecalis. Journ of clin. Micorbiology. 1992; vol.30; (1) p.41-45. | 22. | K H Keddy, K P Klugman, L D Vebowitz. Incidence of high– level gentamicin resistance in enterococci at Johannesburg Hospital. S Afr Med J. 1996: 86: 1273-76. | 23. | Agarwal VA, John VI, Pathak AA. Concomitant high level resistance to penicillin and aminoglycosides in Enterococci at Nagpur, Central India. Indian J Med Microbiol. 1999; 17(2): 85-87. | 24. | Udo EE, Noura A, Phillips OA, Chugh TD. Species prevalence and antibacterial resistance of Enterococci isolates in Kuwait hospitals. J Med Microbiol. 2003; 52: 163-68. | 25. | Bhat KG, Paul C, Bhat MG. High level aminoglycoside resistance in Enterococci isolated from hospitalized patients. Indian J Med Res. 1997 May; 105: 198-99. [ Google Scholar]
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TABLES AND FIGURES |  |
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