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 Table of Contents  
ORIGINAL ARTICLE
Year : 2014  |  Volume : 1  |  Issue : 3  |  Page : 145-149

Evaluation of prevalence and antibiogram of multi drug resistant, extensively drug resistant and pan drug resistant Pseudomonas aeruginosa in patients visiting a tertiary care hospital in central India


1 Department of Microbiology, Sri Aurobindo Institute of Medical Sciences Medical College, Indore, Madhya Pradesh, India
2 Sri Aurobindo Institute of Medical Sciences Medical College, Indore, Madhya Pradesh, India

Date of Web Publication17-Aug-2014

Correspondence Address:
Gunjan Shrivastava
Department of Microbiology, Sri Aurobindo Institute of Medical Sciences Medical College, MR-10 Crossing, Indore-Ujjain Road, Indore - 453 555, Madhya Pradesh
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/2348-3334.138882

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  Abstract 

Introduction: Antibiotic resistance is an increasing concern worldwide, specially in Gram negative bacilli where there is a paucity of new and effective antimicrobial agents. Pseudomonas aeruginosa is inherently resistant to different antimicrobial agents who are responsible for increase in morbidity and mortalities found in all types of patients. The purpose of this study was to evaluate the antibiogram of Multi drug resistant (MDR ), Extremely drug resistance (XDR) and Pan drug resistant (PDR) Pseudomonas aeruginosa. Aims and objectives: To find out the frequency and resistance pattern of MDR, XDR and PDR Pseudomonas aeruginosa. Materials and Methods: This is six month retrospective study, conducted in department of Microbiology a tertiary care hospital and teaching institute. During the study period (July-Dec 2013) routine samples were tested to standard microbiological procedure. Isolates were identified up to species level, Pseudomonas aeruginosa were picked up for further studies. Antibiotic sensitivity testing (AST) was performed by the Kirby-Bauer disc diffusion method, Carbapenemase production was screened by Hodge test and Modified Hodge test, extended spectrum beta lactamse (ESBL) production and AmpC presence were screened by two disc method. According to the sensitivity pattern, the strain was identified as MDR, XDR and PDR Pseudomonas aeruginosa. Results: During the study Pseudomonas aeruginosa were isolated from 198 (21.85%) samples. Among 198, 12 (6.06%) were identified as PDR, 23 (11.6%) were XDR whereas 49 (24.7%) were MDR. Out of 198, 98 (49.49%) were ESBL, 40 (20.20) were Carbapenemase producer and 4 (2.02) were AmpC producer. Conclusions: Though Pseudomonas aeruginosa is inherently resistance to different antimicrobial agents, irrational and inappropriate use of antibiotics is also responsible for the development of resistance to antibiotic. Hence, there is a need to emphasize the "rational drug" to minimize the misuse of antimicrobials. To minimize morbidity and mortalities due to Pseudomonas aeruginosa infection, prior AST is essential to provide specific treatment.

Keywords: Extremely drug resistance, multi drug resistant, pan drug resistant, Pseudomonas aeruginosa


How to cite this article:
Shrivastava G, Bhatambare G S, Patel K B. Evaluation of prevalence and antibiogram of multi drug resistant, extensively drug resistant and pan drug resistant Pseudomonas aeruginosa in patients visiting a tertiary care hospital in central India. CHRISMED J Health Res 2014;1:145-9

How to cite this URL:
Shrivastava G, Bhatambare G S, Patel K B. Evaluation of prevalence and antibiogram of multi drug resistant, extensively drug resistant and pan drug resistant Pseudomonas aeruginosa in patients visiting a tertiary care hospital in central India. CHRISMED J Health Res [serial online] 2014 [cited 2019 Oct 19];1:145-9. Available from: http://www.cjhr.org/text.asp?2014/1/3/145/138882


  Introduction Top


Almost 50 years ago, Pseudomonas aeruginosa was rarely considered as a real pathogen. In the 1970s it was recognized as the microorganism associated with bacteria in the neutropenic host. Nowadays, it is among the most common pathogens involved in nosocomial infections. Hospital reservoirs of the microorganism include respiratory equipment, antiseptics, soaps, sinks, mops and physiotherapy and hydrotherapy pools. [1] It was also noted that Pseudomonas aeruginosa bacteremia is associated with higher mortality than other gram negative bacteremia. [2] Resistance in pathogenic bacteria against antibiotics is a challenge for our clinicians for the management of various infections. [3] Pseudomonas aeruginosa develop resistant to almost all antibiotics by various mechanism like biofilm formation, efflux pumps, resistance genes (SHV, IMP, VIM), aminoglycoside modifying enzyme and mutation in various chromosomal genes [4],[5] and exposure to broad spectrum antibiotics and patient-to-patient spread resistant strains. [6] All those isolates of Pseudomonas aeruginosa, which are resistant to at least one antimicrobial agent in three or more anti-Pseudomonal antimicrobial categories are termed as multi drug resistance (MDR) [7] While, extremely drug resistance (XDR) Pseudomonas aeruginosa are defined as all those isolates which are resistant to at least one antimicrobial agent in six or more anti-Pseudomonal antimicrobial categories. [7] While pan drug resistant (PDR) Pseudomonas aeruginosa refers to those isolates, which are resistant to all pseudomonal antimicrobial categories. [7],[8] On development of new antimicrobial agents, bacteria as well develop resistance against these newer agents and thus these antimicrobial agents are playing an important role in the increased resistance of Pseudomonas aeruginosa. MDR, XDR, PDR strains of Pseudomonas aeruginosa alarming as an important cause of morbidity and mortality. So, clinician should be aware of the frequency and susceptibility pattern of the Pseudomonas aeruginosa in order to manage their patients better.


  Materials and Methods Top


During the study period (July-Dec 2013) routine samples were tested by standard microbiological procedure and isolates were identified by observing the colony characteristics on the blood agar and Macconkey agar plates and biochemical reaction using standard microbiological methods. [9] Isolates were identified up to species level, Pseudomonas aeruginosa were picked up for further study. Antibiotic sensitivity testing (AST) was performed by the Kirby-Bauer disc diffusion method on Muller-Hinton agar. [10] The following antibiotics were tested: amikacin (30 μg), gentamicin (10 μg), piperacillin (100 μg), ticarcillin (75 μg), aztreonam (5 μg), ceftazidime (30 μg), cefepime (30 μg), piperacillin-tazobactum (100/10 μg), colistin (10 μg), imipenem (10 μg) and meropenem (10 μg). (Details of Zone and MIC in [Table 1]) Dehydrated media and antibiotic discs were procured from Hi-Media, Mumbai, India. The control strains used during the study were Pseudomonas aeruginosa ATCC (American Type Culture Collection) 27853. The selection of antibiotics and interpretation of inhibition zone sizes were done according to Clinical Laboratory Standards Institute (CLSI-2013) guidelines. [11] For the detection of carbapenem-hydrolyzing metallo-β-lactamase in Pseudomonas aeruginosa was screened by Hodge test and Modified Hodge test. A lawn culture was prepared on Muller Hinton agar by using an overnight culture of Escherichia coli ATCC 25922 and the turbidity was 0.5 McFarland's standard. The plate was drying for at least 15 minutes and then a disc of 10 μg meropenem was applied at the center of the plate, the test isolate was streaked from the edge of the disc to the periphery of the plate. We tested four isolates per plate. After an overnight incubation at 37°C under aerobic conditions, the clover leaf like indentation between the test streaks near the disc was considered as positive for carbapenemase production. For the detection of bacterial resistance to the penicillins, first, second and third generation cephalosporins, and aztreonam (but not the cephamycins or carbapenems) ESBL detection was done. [11] While performing antibiotic sensitivity testing, ceftazidime plus clavulanic acid (30/10 μg) and cefotaxime plus clavulanic acid (30/10 μg) discs were also included along with ceftazidime (30 μg) and cefotaxime (30 μg) discs on Muller-Hinton agar for ESBL detection. Organism was considered as ESBL producer if there was a ≥ 5 mm increase in the zone diameter of ceftazidime/clavulanic acid disc and that of ceftazidime disc alone and or ≥5 mm increase in the zone diameter of cefotaxime/clavulanic acid disc and that of cefotaxime disc alone. Pseudomonas aeruginosa a known in-house ESBL producer was used as negative and positive controls. [10] Simultaneously AmpC screening by phenotypic method was also done. There are no standardized phenotypic screening methods that are rapidly available in microbiological laboratory for AmpC detection. Isolates showing blunting of ceftazidime or cefotaxime zone of inhibition adjacent to cefoxitin disc or showing reduced susceptibility to either of the above test drugs (ceftazidime or cefotaxime) and cefoxitin were considered as "screen positive" for AmpC detection whereas cefoxitin-cloxacillin double-disc synergy test was used to confirm AmpC production by the Pseudomonas isolates. These isolates were simultaneously confirmed by placing the cefoxitin-cloxacillin (30 μg/200 μg) and cefoxitin (30 μg) at 15 mm apart on Muller-Hinton agar plate pre-inoculated with the test strain and incubated at 35°C for 18-24 hours. A difference in the cefoxitin-cloxacillin inhibition zones minus the cefoxitin alone zones of ≥4 mm was considered indicative for AmpC production. [12]
Table 1: The interpretation criteria and MIC of drug

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According to the sensitivity pattern, the strain were identified as MDR (isolates resistant to at least one antimicrobial agent in three or more anti-Pseudomonal antimicrobial group), XDR (isolates resistant to at least one antimicrobial agent in six or more anti-Pseudomonal antimicrobial group) and PDR (isolates resistant to all anti-Pseudomonal agents). The entire testing was done under strict internal quality control using ATCC (27853) strains. The present study has been approved by ethical committee of our institute.

Inclusion criteria

Samples which showed pure growth of Pseudomonas aeruginosa and isolates which were confirmed in repeat sampling were included in the study.

Exclusion criteria

Mixed growth was excluded in present study.


  Results Top


During the study total of 906 samples were found to be culture positive. Pseudomonas aeruginosa were isolated from 198 (21.85%) samples. Out of 198, 74 (37.37%) were pus, 36 (18.18%) were urine, 32 (16.16%) were BAL, 16 were tip, 12 (6.06%) were blood, 12 (6.06%) were sputum, 10 (5.05%) were tracheal, and 6 (3.03%) were fluid [Table 2]. Among 198, 12 (6.06%) were identified as PDR, 23 (11.6%) were XDR and 49 (24.7%) were MDR. Out of 198, 98 (49.49%) were ESBL producer, 40 (20.20) were carbapenemase producer and 4 (2.02) were AmpC producer. Sensitivity patterns of MDR, XDR and PDR were shown in [Table 3].
Table 2: The prevalence of PDR, XDR, MDR and ESBL, Carbapenemase and AmpC production

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Table 3: The resistance of MDR, XDR, PDR Pseudomonas aeruginosa against various anti-pseudomonal drugs

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  Discussion Top


0Pseudomonas aeruginosa is a well-recognized nosocomial pathogen that can cause severe infection in hospitalized patients. Few treatment options remain for serious infection caused by MDR Pseudomonas aeruginosa. Similarly, option are lacking for XDR and PDR Pseudomonas infection. The isolation rate of Pseudomonas aeruginosa in our set up was 21.85%, which is higher to 18% rate of nosocomial infection reported by God et al.,(2007) [13] While lower incidence reported by Khan et al., (2008) [14] in Pakistan 6.67%. In our study MDR, XDR, PDR Pseudomonas aeruginosa were most commonly isolated from PUS sample followed by Urine and Respiratory samples. In our study MDR Pseudomonas aeruginosa were 24.7%, which were more similar to study conducted in Rawalpindi. [15] Beside that the study conducted in Houston by Tam et al., in 2010 was noted 14%. [16] Farhatullah et al., [17] reported a frequency of 29%, in the burnt patients. While, a prevalence of 45.2% has been reported in India. [18] In another study, Aloush et al., have reported an incidence of 14 MDR Pseudomonas aeruginosa per 10,000 hospital admission. [6] Regarding the antimicrobial susceptibility, in the present study, the highest resistance of MDR was found to be gentamicin followed by ticarcillin, aztreonam, amikacin, pipracillin, meropenem, pipracillin tazobactum which is more similar to the study conducted in 2011. [15] Another study reported that MDR Pseudomonas aeruginosa isolated from blood, highly susceptible to Colistin and 100% resistant to cephalosporine and 21% against aminoglycosids. [19] In another study conducted in Turkey found colistin to be most effective against MDR Pseudomonas aeruginosa. [16],[20] In our study, prevalence of XDR Pseudomonas aeruginosa was 11.6%, which is lower from the another study conducted in 2012. [21] In case of XDR higher resistance was found to ceftazidime followed by amikacin, pipracillin tazobactum. In other study conducted in Singapore hospital, resistance noted for commonly used antibiotics such as Amikacin, Cefepime, Ciprofloxacin and Pipracillin tazobactum was 22.3%. [22] In our study 6.06% isolates were PDR which are resistant to all anti-Pseudomonal drug. In fact the irrational and inappropriate use of antibiotics is responsible for the development of resistance of Pseudomonas aeruginosa to antibiotic monotherapy. Hence, there is a need to emphasize the "rational drug" to minimize the misuse of available antimicrobials. Beside that the regular antimicrobial susceptibility surveillance is essential for area-wise monitoring of the resistance pattern and an effective antibiotic policy should be introduced to presence the effectiveness of antibiotics and better management of patients.

In conclusion, MDR Pseudomonas aeruginosa has commonly been reported in nosocomial infection. Hence based on the present study Colistin and Pipracillin tazobactum has shown highest susceptibility to MDR Pseudomonas aeruginosa. While XDR susceptible to Colistin, but the PDR Pseudomonas aeruginosa was resistant to all anti-pseudomonal drug, hence there is no drug for serious infections caused by PDR Pseudomonas aeruginosa, so that no controlled clinical trials guide therapeutic choice for PDR Pseudomonas aeruginosa, until better antibiotics are being developed now. The selection of empirical antibiotic combination therapy may be difficult due to no common effective antibiotic combination. Hence, we recommend continued surveillance of infection with these microbes, proper antisepsis and urgent treatment with appropriate antibiotics according to the antibiotic sensitivity report for decrease in morbidity and mortality due to Pseudomonas aeruginosa infection.


  Acknowledgment Top


The authors wish to thank the Chairperson and Dean of the institute for providing laboratory facilities and healthy working atmosphere during the study period. The authors are also thankful to the technical staff of the institute for providing necessary helping hand during the endeavor.

 
  References Top

1.Pollack M. Pseudomonas aeruginosa. In: eitor.Principles and Practice of Infectious Diseases. 4 th ed. London: Churchill Livingstone; 1995. p. 1980-2003.  Back to cited text no. 1
    
2.Young LS. The clinical challenge of infections due to pseudomonas aeruginosa. Rev Infect Dis 1984;6:S603-7.  Back to cited text no. 2
    
3.Lister PD, Wolter DJ, Hanson ND. Antibacterial resistant Pseudomonas aeruginosa: Clinical impact and complex regulation of chromosomally encoded resistance mechanisms. Clin Microbiol Rev 2009;22:582-610.  Back to cited text no. 3
    
4.Tam VH, Chang KT, Abdelraouf K, Brioso CG, Ameka M, McCaskey LA, et al. Prevalence, mechanism and susceptibility of multidrug resistant bloodstream isolates of Pseudomonas aeruginosa. Antimicrob Agents Chemother 2010;54:1160-4.  Back to cited text no. 4
    
5.Nwankwo EO, Shuaibu SA. Antibiotic susceptibility pattern of clinical isolates of Pseudomonas aeruginosa in a tertiary health institution in Kano Nigeria. J Med Biomed Sci 2010;17:37-3.  Back to cited text no. 5
    
6.Aloush V, Navon-vanezia S, Siegman-Igra Y, Cabili S, Carmeli Y. Multidrug-resistant Pseudomonas aeruginosa: Risk factors and clinical impact. Antimicrob Agents Chemother 2006;50:43-8.  Back to cited text no. 6
    
7.Magiorakos AP. Multidrug-resistant (MDR), extensively drugresistant (XDR) and pandrug-1 resistant (PDR) bacteria in healthcare settings. Expert proposal for a standardized international terminology; 2011.  Back to cited text no. 7
    
8.ASM Submits Comments on "Multi drug resistant (MDR), extensively drug-resistant (XDR) and Pan drug resistant (PDR) bacteria in healthcare settings. Washington: American Society For Microbiology; 2010. Available online with link http://www.microbemagazine.org/index.php/10-2010-publicaffairs-report/2942-asm-comments-on-ecdc-standardizedinternational-terminology-for-mdr-xdr-and-pdr-bacteria-in-healthcare-settings.  Back to cited text no. 8
    
9.Cheesbrough M. Medical laboratory manual for tropical countries. Vol. 2; Microbiology. 985, England.  Back to cited text no. 9
    
10.Bauer AW, Kirby WM, Sherris JC, Turck M. Antibiotic susceptibility testing by a standardized single disc method. Am J Clin Pathol 1966;44:493-6.  Back to cited text no. 10
    
11.Clinical Laboratory Standards Institute. Performance standard for antimicrobial susceptibility testing. 23 rd Information supplement. NCCLS Document M100-S23; 2013.  Back to cited text no. 11
    
12.Singhal S, Mathur T, Khan S, Upadhyay DJ, Chugh S, Gaind R, et al. Evaluation of methods for AmpC Beta-Lactamase in gram negative clinical isolates from tertiary care hospitals. Indian J Med Microbiol 2005;23:120-4.  Back to cited text no. 12
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13.Gad GF, El-Domany RA, Zaki S, Ashour HM. Characterization of Pseudomonas aeruginosa isolated from clinical and environmental samples in Minia, Egypt: Prevalence, antibiogram and resistance mechanisms. J Antimicrob Chemother 2007;60:1010-7.  Back to cited text no. 13
    
14.Khan JA, Iqbal Z, Rahman SU, Farzana K, Khan A. Prevalence and resistance pattern of Pseudomonas Aeruginosa against various antibiotics. Pak J Pharm Sci 2008;21:311-4.  Back to cited text no. 14
    
15.Gill MM, Usman J, Kaleem F, Hassan A, Khalid A, Anjum R, et al. Frequency and Antibiogram of Multi-drug Resistant Pseudomonas aeruginosa. J Coll Physicians Surg Pak 2011;21:531-4.  Back to cited text no. 15
    
16.Timurkaynak F, Can F, Azap OK, Demirbilek M, Arslan H, Karaman SO. In vitro activities of non-traditional antimicrobials alone or in combination against multidrug-resistant strains of Pseudomonas aeruginosa and Acinetobacter baumannii isolated from intensive care units. Int J Antimicrob Agents 2006;27:224-8.  Back to cited text no. 16
    
17.Ullah F, Malik SA, Ahmed J. Antimicrobial susceptibility and ESBL prevalence in Pseudomonas aeruginosa isolated from burn patients in the North West of Pakistan. Burns 2009;25:1020-5.  Back to cited text no. 17
    
18.Amutha R, Padmakrishnan, Murugan T, Renuga Devi MP. Studies on multidrug resistant Pseudomonas aeruginosa from pediatric population with special reference to extended spectrum beta lactamase. Indian J Sci Technol 2009;2:11-3.  Back to cited text no. 18
    
19.Kirikae T, Mizuguchi Y, Arakawa Y. Investigation of isolation rates of Pseudomonas aeruginosa with and without multidrug resistance in medical facilities and clinical laboratories in Japan. J Antimicrob Chemother 2008;61:612-5.  Back to cited text no. 19
    
20.Bergen PJ, Forrest A, Bulitta JB, Tsuji BT, Sidjabat HE, Paterson DL, et al. Clinically relevant plasma concentrations of colistin in combination with imipenem enhance pharmacodynamic activity against multidrug-resistant Pseudomonas aeruginosa at multiple inocula. Antimicrob Agents Chemother 2011;55:5134-42.  Back to cited text no. 20
    
21.Peña C, Gómez-Zorrilla S, Suarez C, Dominguez MA, Tubau F, Arch O, et al. Extensively drug-resistant Pseudomonas aeruginosa: Risk of bloodstream infection in hospitalized patients. Eur J Clin Microbiol Infect Dis 2012;31:2791-7.  Back to cited text no. 21
    
22.Hsu LY, Tan TY, Tam VH, Kwa A, Fisher DA, Koh TH, Network for Antimicrobial Resistance Surveillance (Singapore). Surveillance and correlation of antibiotic prescription and resistance of Gram-negative bacteria in Singaporean hospitals. Antimicrob Agents Chemother 2010;54:1173-5.  Back to cited text no. 22
    



 
 
    Tables

  [Table 1], [Table 2], [Table 3]


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