Prevalence of Extended Spectrum Beta- Lactamase Producers among Various Clinical Samples in a Tertiary Care Hospital: Kurnool District, India

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1 International Journal of Current Microbiology and Applied Sciences ISSN: Volume Number (17) pp Journal homepage: Original Research Article Prevalence of Extended Spectrum Beta- Lactamase Producers among Various Clinical Samples in a Tertiary Care Hospital: Kurnool District, India Suguneswari Giddi, Sarada Dasari * and C. Suryakumari Department of Microbiology, Viswabharathi Medical College, Penchikalapadu, Kurnool District, A.P., India *Corresponding author A B S T R A C T K e y w o r d s, Escherichia coli,, Susceptibility pattern, Resistance pattern. Article Info Accepted: 1 June 17 Available Online: 1 August 17 Extended spectrum beta lactamase are posing a major obstacle in the therapeutic outcome of patients. They cause enzymatic hydrolysis of the beta lactum ring, resulting in resistance to penicillins and cephalosporins. The rise in results in prolonged treatment which leads to increased hospital stay and financial burden on the patient. It is necessary for all the hospitals to monitor to formulate preventive and therapeutic measures accordingly. The study was conducted in a tertiary care hospital during the period from December 15 to November 1. A total of 5 isolates from Enterobacteriaceae which included Escherichia coli and 1 species isolates were processed. were detected by phenotypic screening and confirmatory methods which included double disc synergy method, Cephalosporin Clavulanate combination disc. Out of Escherichia coli, were (55%) and 3 out of species were (3%). Overall production was 5.% and it was found to be higher in inpatients (55.7%) than outpatients (1.%). All are 1% susceptible to Imipenem and Meropenem and maximum resistance to penicillin and cephalosporins (-9%). pose threat in hospitals; their detection can be made mandatory along with routine antibiogram. Introduction Multidrug resistant bacteria are emerging worldwide in the hospitals and cause treatment failures especially the Enterobacteriaceae group of bacteria. Enterobacteriaceae group of organisms cause various hospital acquired infections like gastrointestinal, urinary tract and pyogenic infections. Among Enterobacteriaceae the most important member that exhibits multiple drug resistance is Escherichia coli (Dinesh Kumar et al., 1). Enterobacteriaceae family possesses plasmid mediated β lactamases and which is the single most important cause of resistance to penicillins 57 and cephalosporins. This resistance is due to mutations in genes like TEM, SHV, and CTX-M. These mutations cause alteration of aminoacid configuration which results in their ability to hydrolyze various beta lactum antibiotics like penicillins, cephalosporins and monobactum (Agarwal, ). Among Enterobacteriaceae the major are Escherichia coli and. are worldwide in distribution and its rate and methods of isolation varies in different areas. are found in clinical samples like urine, blood, sputum, swabs,

2 body fluids, and catheter tips (Kenneth, ). Apart from resistance to broad spectrum cephalosporins and monobactums, also exhibit co-resistance to other groups of antibiotics such as quinolones, cotrimoxazole, and tobramycin, this leads to limitation of available therapeutic options. production is indicated by eight fold reduction in MIC and accentuation of zone of inhibition of third generation cephalosporins in the presence of clavulinic acid (Iraj Alipourfard, 1). High rate of producing organisms associated with fewer therapeutic options lead to longer hospital stay, increased cost, high rate of morbidity and mortality (Folasoge A et al.,, 1). Therefore the present study was conducted to be able to study the prevalence, antibiotic resistance profile of s so that an effective antibiotic strategy can be planned to limit infections due to them. Materials and Methods The present study was conducted in Viswabharathi medical college, a tertiary care hospital in Kurnool, during the period from December 15 to November 1. The isolates were collected from various clinical samples which include urine, blood, sputum, swabs and body fluids. Samples from patients attending both outpatient and inpatient departments were included in the study without applying any selection criteria. Sufficient amount of various clinical samples were collected under aseptic conditions and were inoculated on to blood agar and MacConkey agar. Blood samples were collected under aseptic conditions and inoculated in brain heart infusion agar. They were subcultured on to blood agar and MacConkey agar after overnight incubation at 37ºC for hrs. All the inoculated samples were incubated at 37ºC for hrs and were processed further for identification of the organism by studying colony morphology, motility, and gram staining and biochemical reactions. Isolated organisms were subjected to antimicrobial susceptibility testing by Kirby- Bauer disc diffusion method on Mueller Hinton agar following clinical laboratory standards institute guidelines CLSI guidelines (CLSI, 1). Commercially available antibiotic discs from Hi- media were used. The antibiotic discs used were Ampicillin (1µg), Cephalexin (3µg), Cefoxitin (3 µg), Ceftazidime (3 µg), Cefotaxime (3µg), Ceftriaxone (3µg), Aztreonam (3µg), Imipenem (1µg), Gentamicin (1µg), Amikacin (3µg), Ciprofloxacin (5µg), Norfloxacin (1µg), Nitrofurantoin (3µg), Meropenem (1µg), Amoxyclav (/1µg), Ticarcillin-Clavulinic acid (75/1ug), Piperacillin-Tazobactum (1/1 µg). detection was detected by phenotypic screening and confirmatory tests. Phenotypic screening test Ceftazidime, Aztreonam, Cefotaxime and Ceftriaoxone antibiotic discs were used for antibiotic sensitivity testing as a routine method. The zone of inhibition was measured and it was mm for Ceftazidime, 7mm Aztreonam, 5 Ceftriaxone, and 7mm for Cefotaxime. The above zone sizes indicate suspected. They were confirmed by confirmatory tests. Phenotypic confirmatory tests methods Double disk synergy test (DDS) It is a diffusion test in which 3 µg of Ceftriaxone, Ceftazidime, Cefotaxime and Aztreonam antibiotic discs are subjected to antibiotic susceptibility testing on Mueller Hinton agar with lawn culture of the isolate about 3mm from Amoxicillin Clavulanic 5

3 acid disk. After overnight incubation extension of zone of inhibition of antibiotic disc towards Clavulanic acid containing disc indicates (Dinesh Kumar, 1). Combination disc containing clavulanic acid Cefotaxime and Ceftazidime disc with and without Clavulanic acid were placed at appropriate distance on Mueller Hinton agar plate with isolate and read after overnight incubation. A difference of more than 5mm of zone size between cephalosporin disc and their clavulanate containing disc indicates production (Dinesh Kumar et al., 1). Results and Discussion This study was conducted in a tertiary care hospital at Kurnool district during the period December 15 to November 1. Samples from inpatients and outpatients attending various clinical departments are included in the study. Various clinical samples included in the study were urine, sputum, blood, pus, swabs and fluids. Maximum productions was seen in urine (5.35%) samples followed by swabs (1.59%), blood (.%), fluid (7.7%), sputum (5.51%) (Table 1). Among 5 total Escherichia coli was the dominant organism from urine (9.3%), swab (1.5%), blood (1.1%), fluids (7%), where as in maximum production was seen in sputum samples (5.71%) (Table ). Out of 5 isolates, isolates were Escherichia coli and 1 isolates were spp. Among isolates of Escherichia coli (55%) were and (1.%) were non. Among 1 isolates of species 3 (3%) were and 3 (.57%) were non (Table 3). The overall from total 5 samples were 5.% and non were 19. % (Table ). All the isolates exhibited 1% sensitivity to Imipenem and Meropenem. Among all the isolates.31% showed sensitivity to Pipercillin-Tazobactum, 77.1% to Amikacin and Amoxyclav, followed by 7.37% to Ticarcillin-Clavulanic acid, 5.% to Gentamicin and 5.39% isolates were sensitive to Nitrofurantoin. All the isolates showed maximum resistance (% to 9%) to all - generation Cephalosporins. The resistance pattern for other drugs were 93.7% to Ampicillin, 9.55% to Aztreonam, 7.% to Norfloxacin, and 7.7% isolates were resistant to Ciprofloxacin (Table ). Table.1 Frequency of samples yielding Source Frequency Percentage Urine Swabs Blood Fluids Sputum Total 5 - Extended spectrum betalactamase % 1.59%.% 7.7% 5.51%

4 Table. prevalence among various clinical samples Sample No of isolates Positive Escherichia coli No Percentage No Percentage Urine Swabs Blood Fluids Sputum % 1.5% 1.1% 7% 1.% 1 3.1% 1.75% 1.1% 3% 5.71% Total Extended spectrum betalactamase Urine Swabs Blood Fluids Sputum Escherichia coli E.coli Screening isolates positives Table.3 production in Escherichia coli and Non isolates Screening positives Non (55%) (1.%) (3%) 3 (.5%) E.coli- Escherichia coli, -Extended spectrum betalactamase, Non-- Non Extended spectrum betalactamase Table. and Non in Escherichia coli and Organisms Total isolates & % non- & % Escherichia coli (55%) (1.%) 1 3 (3%) 3 (.57%) Total 5 5 (5.%) 9 (19.%) P- Value <.1 - Extended spectrum betalactamase, Non-- Non Extended spectrum betalactamase Table.5 Number of Enterobacteriaceae isolates in in-patients and out-patients Escherichia coli Specimen IP OP Specimen IP OP Urine (n=5) Swabs (n=5) Sputum (n=3) Blood (n=3) Fluids (n=5) Urine (n=17) Swabs (n=13) Sputum (n=) Blood (n=1) Fluids (n=1) P- Value- <.5 IP- In patient, OP- Outpatient

5 Table. Susceptibility and resistance pattern of Antibiotic Sensitivity Resistance Number Percentage Number Percentage Ampicillin Cephalexin Cefoxitin Ceftazidime Cefotoxime Ceftriaxone Cefaperazone Aztreonam Imipenem Gentamicin Amikacin Ciprofloxacin Norfloxacin Nitrofurantoin Meropenem Amoxyclav Ticarcillin- Clavulinic acid Pipercillin- Tazobactum % 1.53% 19.% 1.5% 9.% 11.1% 1.59% 9.% 1% 5.% 77.1% 9.9% 33.7% 5-39% 1% 77.1% 7.37%.31% % 3.%.3% 5.3% 9.55%.1% 7.% 9.55% 1.73%.3% 7.7% 7.% 9.%.3% 3.% 19.1% - Extended spectrum betalactamase A total of 5 isolates from Enterobacteriaceae were processed for production in our one year study period. Although other members of enterobacteriacae also produce, Escherichia coli and species were taken into consideration. Out of the Escherichia coli isolates (55%) were, 3 (3%) among 1 species were. Overall production was seen in 5 isolates (5.%) which matches with the study by Hima bindu (15) (5.%). production in Escherichia coli and was 55% and 3% respectively in our study which is consistent with studies by Hima bindu (15) (5.%,.3%) and Sasirekha (5.%, 5.1%). Study by Dinesh Kumar et al., (1) showed 55.55% of production in Escherichia coli which is highly similar with our study (55%). production in species in our study was 3% which is similar to the study by Shukla et al., (3.1%) and slightly less than the study done by Baby Padmini et al., () (%). Overall percentage of in India varies between - 75% (Aruna et al., 1). In our study production was found to be high among inpatients (55.7%) than out patients(.%) this can be compared to the study by Dinesh Kumar et al., (1) (IP-.95%, OP-%) and Rinki et al., (1) (IP-71.3%, OP-5.5%). The present study showed highest number of from urine and Escherichia coli was found to be the dominant organism, this observation is in agreement with the study by Folasoge.A.et al.,, In our study highest susceptibility was found for Imipenem and Meropenem (1%) followed by Piperacillin-Tazobactum (%), Amikacin, 1

6 Amoxyclav (77.1%) and Ticarcillin- Clavulanate (7%). Maximum resistance was seen for Ampicillin, all -generations of Cephalosporins and Aztreonam (-9%). This susceptibility and resistance pattern is in accordance with other studies (Pooja Shakya et al., 17; Akila et al., 1 and Dinesh Kumar, 1). The present study was done in a tertiary care hospital at Kurnool district on production among Enterobacteriaceae group of organisms. Escherichia coli emerged as a dominant producer when compared to. All generations of Cephalosporins, and other drugs like Ampicillin, Aztreonam and Ciprofloxacin showed maximum resistance; only Imipenem and Meropenem exhibited 1% susceptibility to. Indiscriminate usage of board spectrum antibiotics as an option of empirical choice of treatment leads to rise of complicating treatment options. Strict implementation of hospital infection control measures along with restricting inadvertent use of cephalosporins may contribute in preventing. References Akila. K., et al., 1. Prevalence of producing species and their in-vitro antimicrobial susceptibility pattern in a tertiary care hospital. IOSR Journal of Dental and Medical Sciences, 5-1. Aruna. K., et al., 1. Prevalence of production among uropathogens in South Mumbai and its Antibiogram pattern. EXCLIJ. 11: Baby Padmini, et al.,. Extended spectrum beta-lactamases in urinary isolates of Escherichia coli and pneumoniae - prevalence and susceptibility pattern in a tertiary care hospital. Indian J Med Microbiol,, Chowdary U., et al.,. Extended spectrum β-alactamases ();An Emerging threat to clinical therapeutics. Indian journal of medical microbiology, (): 75-. CLSI, 1. Performance Standards for Antimicrobial Susceptibility Testing; Twenty-Fourth Informational Supplement. CLSI document M1- S. Wayne, PA: Clinical and Laboratory Standards Institute. Dalela. G., 1. Prevalence of produces among Gram negative bacilli from various clinical isolates in a tertiary care hospital at Jhalawar, Rajasthan, India. Journal of Clinical Diag.Res. ():1-17. Dinesh Kumar. A., et al., 1. Antimicrobial susceptibility profile of extended spectrum betalactamse producing Escherichia coli from various clinical samples. Infectious diseases reserch and treatment. Folasoge, A., et al., 1. A multicenter study of Beta Lactamase resistant Escherichia coli and pneumoniae reveals high level chromosome mediated extended spectrum β Lactamase resistance in Ogun State, Nigeria. Interdisciplinary perspectives on infectious diseases. Hima Bindu. M., et al., 15. Prevalence of production in Escherichia coli and Spp from different clinical samples-a study in a teaching hospital in Telangana, India. International Journal of Current Microbiology and Applied Sciences, (3):3-3. Iraj Alipourfard, et al., 1. Antibiogram of extended spectrum Beta-lactamase producing Escherichia coli and pneumoniae isolated from hospital samples. Bangladesh journal of

7 medical mecrobiology, vol. (1):3-3. Kenneth. T.,. Bacterial Resistance to Antibiotics. Todars Online Textbook of Bacteriology. Pooja Shakya, et al., 17. production among E.coli and Spp causing urinary tract infections: A hospital based study. The Open Microbiology Journal, 11, 3-3. Rinki R., et al., 1. The detection of extended spectrum Beta-Lactamases (s) producing Escherichia coli isolated from clinical samples. International journal of advance research in biological sciences, 3(5):9-15. Sasirekha. B., 13. Prevalence of, Amp C β-lactamases and MRSA among uropathogens and its antibiogram. EXCLIJ, 1:1-. Shukla. R.,. Prevalence of extended spectrum beta-lactamase producing pneumoniae in a tertiary care hospital. Indian J Med Microbiol,, How to cite this article: Suguneswari Giddi, Sarada Dasari and Suryakumari, C. 17. Prevalence of Extended Spectrum Beta- Lactamase Producers among Various Clinical Samples in a Tertiary Care Hospital: Kurnool District, India. Int.J.Curr.Microbiol.App.Sci. (): doi: 3