INTRODUCTION

Carbapenems belong to the β-lactam group of antibiotics. Until recently, carbapenems are considered to be the choice for the therapeutic management of multidrugresistant gram-negative bacterial infections. Currently, the spread of carbapenem-resistant bacteria are in alarming state due to the limited choice in antibiotics for treating infections caused by them (1). Resistance in bacteria to carbapenems is due to the production of carbapenem hydrolyzing enzymes called carbapenemases. These bacteria have the potential to spread rapidly within the hospital environment and also across continents (2). Infections caused by Klebsiella pneumoniae producing carbapenemases are becoming an increasingly significant problem worldwide. Although this enzyme do not represent the first or the sole mechanism of carbapenem resistance, they are remarkable as they are often not detected by routine susceptibility screening and possess an exceptional potential for dissemination

(3). Infections caused by these organisms making the clinicians with serious treatment challenges, due to limited antibiotic options. Efforts are underway to address these varied clinical challenges and have focused on enhanced infection control practices, better screening methods, determination of optimal usage of existing antibiotics, and development of novel antimicrobials. (4).

MATERIALS AND METHODS:

Bacterial isolates:

A total of 20 non repetitive clinical isolates of K. pneumoniae were collected from Saveetha Medical College, Chennai.They were processed for a battery of standard biochemical and were confirmed. Isolates were preserved in semisolid Trypticase soy broth stock and stored at 4^oC until further use.

Antibiotic Sensitivity Testing:

Antibiotic sensitivity test was carried out by Kirby Bauer disk diffusion method with routinely used commercially available antibiotics(HiMedia,Mumbai). These antibiotics include Ampicillin, Amoxycillin, Ceftazidime, Cefotaxime, Amikacin, Gentamicin, Imipenem, Ciprofloxacinas per CLSI 2015 guidelines(5).

Detection of Carbapenemases

Klebsiella pneumoniae isolates were subjected to Modified Hodge test to detect for carbapenemases. An overnight culture suspension of E.coli ATCC 25922 turbidity adjusted to 0.5 McFarland standard was lawn inoculated on the surfaces of the sterile Mueller Hinton agar plates. After brief drying 10µgImipenem discs were placed at the centre of the plates and the test Klebsiella pneumoniae isolates were streaked from the edges of the discs to the periphery of the plates. Then the plates were incubated at 37^oC for overnight. The presence of a 'cloverleaf shaped' zone of inhibition at the junction point between Imipenem discs and test strains were identified as carbapenemases positive isolates(6).

RESULT:

Sample Wise Distribution of Clinical Isolates of Klebsiella pneumoniae: Of the 20 clinical isolates of Klebsiella pneumoniae, 12/20(60%) were from urine, 4/20(20%) from stool, 3/20(15%) and 1/20(5%) were from the wound swab and pus respectively.

Antibiotic Susceptibility Testing

Increased percentage of isolates were showing resistance to cephalosporins and other group of antibiotics(80-100%). We found very less number of isolates were sensitive to imipenem (20%) which is considered to be a most potential drugs The detailed resistant pattern of Klebsiella isolates were showed in table 1.

Table 1 Results of antibiotic susceptibility patterns of Klebsiella pneumoniae:

Antibiotics	Sensitivity (%)	Intermediate (%)	Resistant(%)
Ampicillin	5	0	95
Amoxicillin	5	0	95
Ceftazidime	5	0	95
Cefotaxime	0	0	100
Amikacin	0	0	100
Gentamicin	15	5	80
Imipenem	20	0	80
Ciprofloxacin	0	0	100

Carbapenemase Production by Klebsiella pneumoniae:

None of our isolates where found to show positivity for carbapenemase production by modified Hodge test

Discussion:

Reports indicate that carbapenemases producing Enterobacteriaceae isolates seem to be increasing in number in the last few years (1). Study conducted by Nagaraj in 2012 from Bangalore reported a a high incidence of bla_NDM among E. coli and K. pneumoniae by PCR (7). Among the carbapenem- resistant Entero-bacteriaceae isolates, NDM-1 was detected in 75% (27/36) of K. pneumoniae and 66% (10/15) of E. coli by PCR. Deshpande et al. reported a similar finding from a tertiary care hospital in Mumbai, in which majority of bla_NDM producing isolates were K. pneumoniae and E. coli (8). However, we seldom tried PCR analysis in our study.

Nagaraj and his colleagues reported that 33% of his Enterobacteriaceae clinical isolates obtained from different clinical specimens were showed positivity for carbapenemase production by Modified Hodge test (7). While in our present study we did not find any positive for carbapenemase production by this method.

CONCLUSION:

The present study was designed as a pilot study; so, the sample size was small and the isolates were consecutive and not random. From this study we did not find any positive carbapenemase producing Klebsiella pneumoniae. For confirming the possibility of nosocomial spread, further study is required with higher sample size, more demographical characteristics, hospitalisation data of patients and molecular methods to determine clonality.

Acknowledgment:

We thank Dr. Kalyani, Professor and Head of the Department of Microbiology, Saveetha Medical College, Chennai for kindly providing the clinical isolates to carry out our research work.

REFERENCE:

1. Walsh TR. Emerging carbapenemases: A global perspective. Int J Antimicrob Agents 2010;36 Suppl 3:S8-14.

2. Cornaglia G, Rossolini GM. The emerging threat of acquired carbapenemases in Gram-negative bacteria. Clin Microbiol Infect 2010;16:99-101.

3. Paterson DL, Bonomo RA. Extended-spectrum beta-lactamases: a clinical update. Clinical Microbiology Reviews. 2005; 18:657–686.

4. Ryan S. Arnold, Kerri A. Thom, Saarika Sharma, Michael Phillips, J. Kristie Johnson, Daniel J. Morgan. Emergence of Klebsiella pneumoniae Carbapenemase (KPC) Producing Bacteria. South Med 2011; 104:40-45.

5. Clinical Laboratory Standards Institution: Performance standards for antimicrobial susceptibility testing. In NCCLS approved standard M2-A8. Wayne, PA USA: CLSI; 2015.

6. Lee K, Chong Y, Shin HB, Kim JJ, Young D, Yum JH. Modified Hodge and EDTA disk synergy tests to screen metallo beta lactamase producing strains of pseudomonas and Acinetobacter species. Clin Microbial Infect 2001; 7:88-91.

7. Nagaraj S, Chandran SP, Shamanna P, Macaden R. Carbapenem resistance among Escherichia coli and Klebsiella pneumoniae in a tertiary care hospital in south India. Indian J Med Microbiol 2012; 30:93-95.

8. Deshpande P, Rodrigues C, Shetty A, Kapadia F, Hedge A, Soman R. New Delhi Metallo-betalactamase (NDM-1) in Enterobacteriaceae: Treatment options with carbapenems compromised. J Assoc Physicians India. 2010;58:147–9.

Received on 10.06.2016 Modified on 22.06.2016

Research J. Pharm. and Tech 2016; 9(10):1585-1587.

DOI: 10.5958/0974-360X.2016.00312.7