Introduction:
Pseudomonas aeruginosa isolates are responsible for outbreaks of nosocomial infections in the world. Pseudomonas aeruginosa producing Metallo ß Lactamases (MBLS) was first reported from Japan in 1991 and since then has been described from various parts of the world including Asia, Europe, Australia, South America and North America. (1) A five years longitudinal study involving many centers from Latin America indicated that year after year, Ps. aeruginosa resistance has continually risen to the point where approximately 40% are resistance to “antipseudomonal” drugs including carbapenems. While the advent of carbapenems in the 1980s heralded a new treatment option for serious bacterial infections caused by cephalosporin and penicillin resistant bacteria, carbapenems resistance can now be observed in Enterobacteriaceae and Acinetobacter spp. and is becoming common place in Ps. aeruginosa.[2,3]
Acquired Metallo –ß- Lactamases (MBLs) have recently emerged as one of the most worrisome resistance mechanisms owing to their capacity to hydrolyze with the exception of aztreonam, all ß lactams including carbapenems and also because their genes are carried on highly mobile elements, allowing easy dissemination.[4,5] The prevalence of imipenem resistance to Ps. aeruginosa has been increasing worldwide. Resistance to carbapenem is due to impermeability via the loss of the Opr D porin, the up regulation of an active efflux pump system of the cytoplasmic membrane or the production of metallo-ß- lactamases (MBLs).
MBLS also represent a clinical threat due to their unrivalled spectrum of activity and their resistance to therapeutic serine ß Lactamases inhibitors. The fact that MBL and aminoglycoside resistance genes are genetically linked merely compounds this problem. The problem of an appropriate treatment regimen is also amplified by the lack of new antimicrobials that will possess broad spectrum potency against clinically significant Ps. aeruginosa. It is necessary to identify the prevalence of these strains in hospitals and to characterize their epidemiology to control the spread of these strains and to determine suitable prevention and treatment policies. Bearing in mind this massive problem of MBL producing Pseudomonas aeruginosa the present study was carried out.
Materials and Methods
A total of 608 Pseudomonas aeruginosa isolates were isolated during the study period. Out of the total 608 Pseudomonas aeruginosa isolates 91 were screen positive that is resistant to Imipenem, they were considered to be putative MBL producers. MBL producers were confirmed by different phenotypic tests i.e. Imipenem –EDTA combined disk Test, Imipenem –EDTA double –disk synergy Test and then later on EDTA disk potentiation test using 4 cephalosporins and E-test. Among 91 putative MBL producers, 81(13.32%) were confirmed as MBL producers by Imipenem-EDTA combined disk test and imipenem –EDTA double disk synergy test and later on EDTA disk potentiation test using four cephalosporins and Epsilometer test. Identification of microorganism was done by standard laboratory technique.3 This figure of 81 confirmed MBL producers is from total sample size of 608. Hence the prevalence rate of MBL producing Pseudomonas aeruginosa is 81/608 i.e.13.32%. 55 (67.90%) of these were from inpatient units and 26 (32.08%) were from outpatient units. Different clinical specimens collected from patients admitted under different clinical disciplines of Krishna Hospital and Medical Research Centre, Karad during 15 Oct 2008 to 15 March 2012, were used in the study.
Metallo Beta Lactamase Detection:
]MBL production was carried out in Imipenem –resistant isolates [1], MBL detection tests were done by following methods using Ps. aeruginosa ATCC 27853 (Hi-media) as a negative control strain.[6] An inhouse Ps. aeruginosa strain which was repeatedly MBL positive by Imipenem – EDTA combined disc test and Imipenem –EDTA double disc synergy test was used as positive control strain.[7,8]
Imipenem – EDTA combined disc test:
The IMP-EDTA combined disk test was performed as described by Yong et al.[9] Pseudomonas aeruginosa ATCC 27853 (Hi-media) was used as the control strain. The test organisms were inoculated on Muller Hinton agar plates as per CLSI guidelines. A 0.5 M EDTA solution was prepared by dissolving 18.61 g of EDTA in 100 ml of distilled water and adjusting its PH 8.0 by using NaOH. The mixture was sterilized by autoclaving. The two 10 µg Imipenem disks were placed on Muller Hinton agar plates, and 5 µl solution of ethylene diamine tetra acetic acid (EDTA) was added to one of Imipenem disk to obtain desired concentration of EDTA i.e.750 µg. And then plates were incubated at 370C for overnight. The inhibition zones of Imipenem and Imipenem plus EDTA were compared after incubation.[6] The increase in zone of inhibition for Imipenem plus EDTA was = 7mm than Imipenem alone then the test were considered as a positive for MBL production.
Imipenem – EDTA double disc synergy test (DDST):
The Imipenem – EDTA double disk synergy test was performed as described by Lee. et. al.[10] The test organisms were inoculated on Muller Hinton agar as recommended by CLSI guideline. An Imipenem disc (10µg) was placed 20mm centre to centre from blank disc containing 5 µl of 0.5M EDTA (750µg). Then plates were incubated at 370C for overnight.[6] Enhancement of zone inhibition in the area between Imipenem and EDTA on blank disk in comparison with the zone of inhibition on the far side of drug was interpreted as a positive result.
EDTA disk potentiation test using Ceftazidime, Ceftizoxime, Cefepime, and Cefotaxime:
Test organisms were inoculated on Muller Hinton Agar plate as per procedure used in standard disk diffusion test recommended by CLSI guidelines. A filter paper blank disk (Whatmann filter paper no.-2) was placed at centre and Ceftazidime (30µg), Ceftizoxime (30µg), Cefepime (30µg), Cefotaxime (30µg) were placed 25mm distance from centre to centre from blank disc, then 5µl solution of 0.5M EDTA solution was added to blank disc and plates were incubated at 370C for overnight. Then results were recorded.[11] Following incubation period for 16-18 hours at 370C a clear extension of zone of inhibition in the area between EDTA disc and any one of the cephalosporin disc in comparison with the zone of inhibition on the far side of drug was interpreted as positive for MBL production.
MBL E- Test:
The E- test of MBL strip containing a double sided seven dilution range of Imipenem (4-256 µg/ml) and IMP (1-64 µg/ml) in combination with a fixed concentration of EDTA used for MBL detection. MIC ratio of Imipenem/Imipenem + EDTA of >8, or reduction of Imipenem MIC by =3 log 2 dilutions in the presence of EDTA or appearance of a phantom zone indicates MBL production.
The strains which were positive for Imipenem – EDTA combined disk test, Imipenem – EDTA double disk synergy test (DDST), EDTA disk potentiation test and MBL- E Test were considered to be MBL producers. And these isolates were further processed for confirmation of transfer of drug resistance by conjugation.[6,12,13]
Transfer of resistance by conjugation:
As the recipient strain was E. coli J53AZR, selection was made on MacConkeys agar containing sodium azide 200 µg/ml Plus each drug (8µg/ml) separately to which the donor strain was resistant (i.e. sodium azide + Cefotaxime, sodium azide + Ceftriaxone, sodium azide + Ceftazidime.)
The donor strain was grown in nutrient broth to the late log phase at 370C until it reached 2×108 CFU, while recipient strain was grown in nutrient broth to the late log phase at 370C until it reached 5×108 CFU. One part of donor and nine parts of recipient cultures were mixed together and then the mixture was incubated with shaking at 370C; similarly another set of same mixture was incubated with shaking at room temperature for two hours.[14]
Loopful mixture from both sets of mixture was then streak inoculated on half of the selection plate and donor and recipient strains were also inoculated on MacConkeys agar plate, which served as contro.
The transfer of resistance factor from MBL positive Ps. aeruginosa (donor strain) to sodium azide resistant E. coli (recipient strain) i.e. (E. coli J53AZR) was done by conjugation.[15-17]
Results
In our study out of 608 isolates of Pseudomonas aeruginosa 91/608 (14.96%) were resistant to imipenem. We have confirmed that 81/608(13.32%) isolates are MBL producers by imipenem -EDTA combined disk test, Imipenem – EDTA double disc synergy test (DDST). Among these 55(67.90%) were from inpatient units and 26(32.08%) were from outpatient units.
Table 1: Distribution of MBL and non MBL producing isolates of Pesudomonas aeruginosa in different specimens. |
| Pseudomonas aeruginosa (N=91) |
Specimens |
MBLs producers |
Non-MBLs producers |
Total |
Urine |
17 (20.98%) |
02(20%) |
19 |
Pus |
38 (46.51%) |
06(60%) |
44 |
Sputum |
06 (07.40%) |
00 |
06 |
Blood/ Bone marrow |
02 (02.46%) |
00 |
02 |
Body fluids |
05 (06.17%) |
00 |
05 |
Others |
13 (16.04%) |
02(20%) |
15 |
Total |
81 |
10 |
91 |
Out of these 81 MBL producing clinical isolates of Pseudomonas aeruginosa, 38(46.51%) were from pus, 17(20.98%) were from urine, 6(07.40%) were from sputum, 5(06.17%) were from body fluids and 2(02.46%) were from Blood/Bone marrow. Likewise MBL producing Ps. aeruginosa were more prevalent in surgery ward i.e. 18 isolates (32.72%) followed by medicine ICU 11(20.00%), Orthopaedic and Medicine 10(18.18%).
Majority of MBL producing isolates of Pseudomonas aeruginosa had a high MIC in the range of 64 = 64 µg/ml for Ceftriaxone and 75/81 (92.59%) isolates had a MIC in the range of 16 -= 64 µg/ml for Ceftazidime. All 81/81 (100.00%) isolates had a MIC in the range of 64-=512 µg/ml for Cefotaxime, 80/81 (98.76%) isolates had a MIC in the range 128-= 512 µg/ml for Piperacillin, 80/81 (98.76%) had a MIC in the range of 4- = 8 µg/ml for ciprofloxacin, 80/81 (98.76%) had a MIC in the range of 16- =256 µg/ml for imipenem.
In India prevalence of MBLs range from 8-87% with a recent study reporting 41% occurrence.[18] In our study the prevalence rate of MBL producing Ps. aeruginosa was 13.32% which was similar to studies conducted by Navneeth BV et al (12%)[19], Rajput A et al (12%)[20], Hemalata et al (14%)[21], Attal RO et al. (11.04%)[22] respectively from various parts of India.
Table 2: Comparison of phenotypic tests used for detection of MBL producing Ps. aeruginosa |
Organism |
Total number of isolates |
Phenotypic tests |
Pseudomonas aeruginosa |
91 |
Imipenem –EDTA combined disk Test; |
Imipenem –EDTA double –disk synergy Test; |
EDTA disk potentiation test using 4 cephalosporins |
Epsilometer Test (E-test) |
81(89.01%) |
81(89.01%) |
76 (83.51%) |
63(69.23%) |
Phenotypic tests i.e., Imipenem –EDTA combined disc test (CDT) and Imipenem –EDTA double disc synergy test (DDST) could detect equal number of MBL producing strains of Pseudomonas aeruginosa, i.e., 89.01%, where as EDTA disc potentiation test using Ceftazidime, Ceftizoxime, Cefepime, Cefotaxime detected 83.51% and Epsilometer test (E-test) detected 69.23%./P>
Table 3: Resistance transfer experiments of MBL producing isolates of Pseudomonas aeruginosa |
| Organism |
No. of isolates |
No. of transfer |
Positive percentage of transfer |
Ps. aeruginosa |
81 |
62 |
76.54% |
Of MBL producing strains, 62/81 (76.54%) could transfer resistance of third generation cephalosporins and antibiotics to the recipient strain. The frequency of transfer was more at 37% than at room temperature. Predisposing risk factor associated with MBL producing Pseudomonas aeruginosa were mainly prolonged hospital stay and the use of Foleys catheter.
The emergence of acquired MBLs among Pseudomonas aeruginosa represents an epidemiological risk for at least two reasons. Firstly MBLs confer resistance not only to carbapenems but to virtually all ß lactams and are frequently associated with resistance to aminoglycosides, and secondly genes encoding to MBL enzymes are most commonly carried on mobile genetic elements (integrons, plasmids, transposons) that can spread horizontally among unrelated strains.[23]
In the present study the prevalence rate of MBL producing strains of Pseudomonas aeruginosa was 81/608 i.e., 13.32%.
All the isolates were 100% sensitive to Aztreonam; they were also sensitive to polymyxin B 96.70% and Colistin 87.91%. Transfer of resistance could be demonstrated from donor to recipient strain. Majority of our MBL isolates are from indoor patients. This is a cause of concern as the percentage of transfer i.e., 76.54% is quite high and according to our literature search this is one of the largest transfers done. Discussion:
Since the widespread use of carbapenem in the hospitals, carbapenem –resistance has been detected increasingly worldwide. After the discovery of penicillin and sulfonamides patients were treated empirically and the organisms were mostly susceptible. After the emergence of resistance occurred, it required the development of new beta lactam antibiotics; with new class of antibiotics, a new beta lactamase emerged that caused resistance to that class of drug. Carbapenem resistance in Pseudomonas spp. is an emerging problem and is a cause of concern as many nosocomial Pseudomonas are detected to be resistant to most of other antibiotics.(23)
MBLs have been identified from clinical isolates worldwide with increasing frequency over the past few years, and strains producing these enzymes have been responsible for prolonged nosocomial outbreaks that were accompanied by serious infections. The occurrence of an MBL positive isolate in a hospital setting poses a therapeutic problem, as well as a serious concern for infection control management. The identification and reporting of MBL producing Ps. aeruginosa will aid infection control practitioners in the spread of this multidrug resistant isolate. (1)
The clinical samples included in the present study were from urine, pus, sputum, blood/bone marrow and body fluids. All the methods for detection of MBL producing bacterial isolates depend on the principle, that MBLs are affected by removal of zinc from their active site. Still, no single screening method has been found to be perfect. Currently, there is no Clinical Laboratory Standard Institute (CLSI) recommended method available. Also, no standard method is recommended by any other international committee for the detection of MBL producers.[22]
A total of 608 Ps. aeruginosa clinical samples collected during the three year period of the study, 91/608 (14.97%) were resistant to imipenem were considered to be putative MBLs producers, out of which 81/608 (13.32%) isolates were confirmed for positive MBL activity. These isolates were confirmed by using Imipenem – EDTA combined disk test, Imipenem – EDTA double disk synergy test, and also by EDTA disk potentiation test using Ceftazidime, Ceftizoxime, Cefepime, Cefotaxime, and MBL E- Test were done. In the present study 91(14.97%) were resistant to imipenem, out of which 81 were confirmed MBL positive by Imipenem –EDTA combined disk Test(CDT) and Imipenem –EDTA double –disk synergy Test (DDST), hence the prevalence rate of MBL producing isolates of Pseudomonas aeruginosa during the period 15 Oct. 2008-15 Mar 2012 is 81/608(13.32%).
In the present study our isolates were less resistant to imipenem i.e. 14.97% (91/608). In other studies the percentage of imipenem resistance ranges from 15.71% to 98.1%. Pandya Y. et al [24] reported Imipenem resistant to be 35/283 (12.3%) which correlates well with the present study.
In the present study, all MBL producing isolates were found to be 100% multidrug resistant. All isolates were resistant to Cephaloridine and Imipenem. Out of 81 MBL producing Ps. aeruginosa 100.00% were resistant to Ceftazidime, 98.76%, 97.53%, 95.06%, and 97.53% were resistant to Cefpodoxime, Cefotaxime, Ceftriaxone, and Cefuroxime. Similarly multidrug resistance was recorded by Bijayini B et al in 2008; 70% isolates of Pseudomonas aeruginosa were resistant to ceftazidime, 75% to Piperacillin, 59% to Piperacillin /tazobactam, 89% to Ticarcillin/ clavulanic acid, 82% to cefoperazone, 74% to amikacin, 81% to cefepime, 71% to levofloxacin, 79% to ciprofloxacin, and 69% to aztreonam by disc diffusion method.[25]
In the present study the MIC of Ceftriaxone, Ceftazidime, Cefotaxime, Piperacillin, Ciprofloxacin was determined against MBL producing and non MBL producing isolates of Pseudomonas aeruginosa, by agar dilution method. MIC of Imipenem was determined by using E- test as Imipenem powder was not available commercially. The MBL producing isolates of Pseudomonas aeruginosa 80/81 ( 98.76%) had a MIC in the range of 64-=64 µg/ml for Ceftriaxone. 75/81 (92.59%) isolates had a MIC in the range of 16-=128 µg/ml for Ceftazidime. All 81/81(100.00%) isolates had a MIC in the range of 64-512 µg/ml for Cefotaxime, 80/81(98.76%) isolates had a MIC in range 256-=512µg/ml for Piperacillin, 80/81( 98.76%) had a MIC in the range of 4-=8 µg/ml for Ciprofloxacin and 80/81(98.76%) had a MIC in the range of 16-=256 µg/ml for Imipenem.
In the present study MIC value for isolates are quite high as compared to other studies. It can be noted that our MIC values are high in comparison to studies carried out abroad. It is also interesting to note that a MIC value of non MBL producers was quite high for most of the antibiotics.
There are only very few reports with reference to resistance transfer experiments in respect to Metallo beta lactamase producing isolates. Among the various modes of gene transfer conjugation is most common mode of resistance gene transfer. The transfer of a plasmid carrying a metallo –ß- lactamase gene suggests the possibilities of clinical spread of plasmid –encoded metallo beta lactamases by cell to cell contact because metallo beta lactamases confer resistance not only to carbapenems but also to other ß lactams except aztreonam (monobactams), antibiotics are frequently ineffective against organisms carrying this enzyme.[13] Atul Khajuria et al., in 2013 from Pune successfully transferred plasmid carrying blaNDM-1 from Pseudomonas aeruginosa to E. coli J53 recipient strain, only in four strains.[26]
In the present study, 62/81 (76.54%) of MBLs producing Ps. aeruginosa isolates could transfer the resistance of Imipenem, Ceftriaxon, Cefotaxime and Ceftazidime to the recipient E.coli J53AZR strain at 370C. By far this is the largest number and percentage of transfer of drug resistance carried out in India as per our literature search.
Conclusion:
MBLs have become a wide spread serious problem and several aspects of them are worrying. MBLs compromise the activity of antibiotics creating therapeutic difficulties with a significant impact on the outcome of patients. In the hospital environment plasmids could be transferred easily between patients through healthcare workers due to hand carriage and of selection pressure.
The early detection of MBL producing Pseudomonas aeruginosa may help in appropriate antimicrobial therapy and avoid the development and dissemination of these multidrug resistance strains. Hence all Pseudomonas aeruginosa isolates resistant to imipenem should be screened for MBL production. Imipenem –EDTA combined disc test. (CDT) and Imipenem –EDTA double disc synergy test (DDST) which are easy to perform and cheap, should be introduced in every clinical microbiology laboratory to detect MBLs in Pseudomonas aeruginosa and to improve disease management. Also framing of rationale antibiotic policy will go a long way in helping the cause.
References:
- Pitout JDD, Gregson DB, Poirel L, McClure J, Phillipze, Church DL. Detection of Pseudomonas aeruginosa producing Metallo –ß- Lactamases in a Large Centralized Laboratory. Journal of Clinical Microbiology, July 2005;43(7):3129-3135. Available at http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1169086/
- Toleman MA, Simm AM, Murphy TA, et.al. Molecular characterization of SPM -1, a novel metallo-ß–Lactamases isolated in Latin America: report from the SENTRY antimicrobial surveillance programme. Journal of Antimicrobial Chemotherapy. 2002;50:673-679.
- Walsh TR, Toleman MA, Poirel L, Nordmann P. Metallo –ß- Lactamases: the Quiet before the storm? Clinical Microbiology Reviews. Apr 2005;18(2):306-325. Available at http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1082798/
- Tsakris A, Poulou A, Pournaras S, et.al. A simple phenotypic method for the differentiation of metallo ß lactamases and class A KPC carbapenemases in Enterobacteriaceae clinical isolates. J Antimicrob Chemother. 2010;65:1664-1671.
- Singla P, Sikka R, Deep A, Chaudhary U. Phenotypic detection and prevalence of Metallo ß- Lactamases (MBLs) in carbapenem resistant isolates of Acinetobacter species at a tertiary care hospital in North India. Int. J. Pharm. Med. & Bio. Sc. 2013;2(1):85-91. Available at http://www.ijpmbs.com/ijpmbsadmin/upload/ijpmbs_50ea9f98db1ae.pdf
- Nagaveni S, Rajeshwari H, Oli AK, Patil SA, Chandrakanth. Incidence of metallo beta –lactamase producing Pseudomonas aeruginosa in clinical samples. The Bioscan. 2010;5(20):251 -253.
- Babu KVY, Kumari A, Kumar A., Raghu Kumar K.G. Role of imipenem –resistant metallo -beta –lactamase positive Pseudomonas aeruginosa carriers in nosocomial infections. Journal of Natural Science, Biology and Medicine January 2013;4(1):181-186. Available at http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3633274/
- Behera B, Mathur P, Das A, Kapil A, Sharma V. An Evaluation of Four Different Phenotypic Techniques for Detection of Metallo ß-Lactamases producing Pseudomonas aeruginosa. Indian Journal Of Medical Microbiology. 2008;26(3):233-37.
- Gupta E, Mohanty S, Sooel S, Dhawan B, Das BK, Kapil A. Emerging resistance to carbapenems in a tertiary care hospital in North India. Indian J Med Res. July 2006;124:95-98.
- Marchiarao P, Maria A, Mussi, et.al. Sensitivity EDTA-Based Microbiological assay for detection of Metallo beta lactamases in Nonfermentative Gram –Negative Bacteria. Journal of Clinical Microbiology. Nov 2005;43(11):5648-5652. Available at http://jcm.asm.org/content/43/11/5648.long
- Franiczex R, Dolna I, Krzyzanowska B. Transferable resistance to different antimicrobial due to CTX-M-type ß-lactamases among Citrobacter Freundii, Serratia marcescense, and Enterobacter spp. clinical isolates. Adv Clin Exp Med. 2007;16(4):493-500.
- Franiczex R, Dolna I, Krzyzanowska B, Szufnarowski K., Kowalska-Krochmal B. Conjugative Transfer of multiresistance plasmids from ESBL-positive Escherichia coli and Klebsiella spp. clinical isolates to Escherichia coli Strain K12 C600. Adv Clin Exp Med 2007;16(2):239-247.
- Ito H, Arakawa Y, Ohsuka S, Wachrotayanku R, Kato N, Ohta M. Plasmid mediated disseminstion of the metallo –B- lactamase gene bla IMP among clinically isolated strains of Serratia marcescens. Antimicrobial Agents and Chemotherapy. Apr 1995;39(4):824-829. Available at http://aac.asm.org/content/39/4/824.long
- Yano H, Kuga A, Okamoto R, Kitasato H, Kobayashi T, Inoue M. Plasmid-encoded Metallo-ß-Lactamase (IMP-6 conferring resistance to carbapenems, especially meropenem. Antimicrobial Agents and Chemotherapy. May 2001;45(5):1343 -1348. Available at http://aac.asm.org/content/45/5/1343.long
- Rit K, Chakraborty B, Dey R, Chakraborty P, Naha A. Prevalence of Pseudomonas aeruginosa and Acinitobacter spp producing Metallo-B- lactamase in a tertiary care hospital. J. Dr NTR Univ Health Sci 2013;2:18-21.
- Navaneeth BV, Sridharan D, Sahay D, Belwadi MRS. A preliminary study on metallo beta lactamase producing Pseudomonas aeruginosa in hospitalized patients. Indian J Med Res. 2002 Dec;116:264-7.
- Rajput A, Prajapati B, Chauhan B, Shah A, Trivedi T, Kadam M. Proevalence of Metallo- betalactamases (MBL) producing Pseudomonas aeruginosa in tertiary care hospital. Indian Journal of Basic and Applied Medical Research. September 2012;1(4):304-308.
- Hemalatha V, Sekar U, Kamat V. Detection of Metallo beta lactamase producing Pseudomonas aeruginosa in hospitalized patients. Indian J Med Res. August 2005;122:148-152.
- Attal RO, Basak S, Mallick SK, Bose S. Metallo beta lactamase producing Pseudomonas aeruginosa: an emerging therat to clinicians. Journal of Clinical and Diagnostic Research. Aug 2010;4:2691-2696.
- Hammami S, Ben Boubaker IB, Ghozzi R, Saidani M, Amine S, Redjeb B. Nosocomial outbreak of imipenem-resistant Pseudomonas aeruginosa producing VIM-2 metallo-B-lactamase in a kidney transplantation unit. Diagnostic Pathology. 2011;6:106.
- Pandya NP, Prajapati SB, Mehta SJ, Kikani KM, Joshi PJ. Evaluation of various methods for detection of metallo B- lactamase (MBL) production in gram negative bacilli. Int J Biol Med Res. 2011;2(3):775-777.
- Franklin C, Liolios L,Peleg AY. Phenotypic detection of carbapenem susceptible metallo-B-lactamase-producing gram –negative bacilli in clinical laboratory. J Clin. Microbiol. 2006;44(9):3139.
- Bijayini B, Anupam D, Purva M, Arti K. High prevalence of carbapenem resistant Pseudomonas aeruginosa at a tertiary care centre of north India. Are we under-reporting? Indian J. Med. Res. Sep 2008;128(3):324-5.
- Khajuria A, Praharaj AK, Kumar M, Grover N. Emergence of NDM-1 in the clinical isolates of Pseudomonas aeruginosa in India. Journal of Clinical Diagnostic Research. Jul 2013;7(7):1328-1331.
- Qu TT, Zhang J, Wang J et al. Evaluation of phenotypic tests for detection of metallo-B-Lactamase-producing Pseudomonas aeruginosa strains in China. J. Clin. Microbiol. 2009,47(4):1136-1142. Available at http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2668318/
- Picao RC, Andrade SS, Nicoletti AG, et al. Metallo-B –lactamase detection: comparative evaluation of double –disk synergy versus combined disk tests for IMP-,GIM-, SIM-, SPM-,or VIM-producing isolates. Journal of Clinical Microbiology. 2008;46(6):2028-37. Available at http://jcm.asm.org/content/46/6/2028.long
|
