Bacteriological profile and sensitivity pattern of microorganisms causing Urinary Tract Infection at a tertiary care center in eastern Uttar Pradesh

Introduction: Urinary tract infection (UTI) is one of the most common infections worldwide and the pattern of antimicrobial susceptibility varies widely in different geographical regions depending on the antibiotic policies. Aim: To know the bacteriological profile of UTIs and the antibiogram of uropathogens in eastern Uttar Pradesh. Material and Methods: This study was prospective and conducted at tertiary care center in one year study duration (2015). All the patients clinically suspected of having UTI were enrolled, culture and sensitivity was performed as per standard protocol, irrespective of age, sex, indoor/outdoor and associated illnesses. Results: In a total of 2217 patients, 967 (43.61 %) were positive for uropathogen by culture, and positivity in female was high (46.48%) as compared to male (38.46%). Escherichia coli was the most common 346 (35.80 %) uropathogen followed by Klebsiella pneumoniae 183 (18.92%) and Enterococcus species 125 (12.92%). Gram negative isolates were most sensitive to imipenem (92%), ceftazidime-clavulanic acid (71%), piperacillin-tazobactam (68%), amikacin (60%), followed by nitrofurantoin (56%). Gram positive isolates were most sensitive to vancomycin (91%) followed by linezolid (80%) and amoxycillin-clavulanic acid (45 %). Conclusion: UTI is a very common problem and rate of antibiotic resistance is relatively high. Imipenem, ceftazidimeclavulanic acid, piperacillin-tazobactam, amikacin, vancomycin and linezolid, were found sensitive against isolated uropathogens.


Introduction
Urinary tract infections (UTIs) are the most common bacterial infection encountered in tertiary care settings. [1] Urine samples are the largest single category of specimens received by most microbiology laboratories, but the majority of the urine culture yield clinically insignificant results. [2] The diagnosis of UTI is primarily based on signs and symptoms rather than isolated laboratory findings; importantly, bacteriuria is not a disease thus, the collection and interpretation of urine cultures should be based on the clinical scenario. [3] Generally, women with acute uncomplicated cystitis culture are not recommended. However, for individuals with acute pyelonephritis or complicated UTI it is important to obtain a urine culture in order to find the appropriate antimicrobial regimen, prior to start empiric therapy. [4] Antimicrobial prescription should be prudent and rational. The choice of antimicrobial agents should be preferably based on the patient's allergy history, antibiogram pattern, availability, cost and compliance. [5] Overuse of antimicrobial in clinical situations where they are not necessary or in prolonged courses of therapy when shorter durations are effective, are responsible for causing antibiotic resistance and it renders a major public health problem worldwide. [6] Thus, the purpose of this study is to know the antibiogram of uropathogens and to guide the patients for appropriate antibiotics.

Material and Methods
The present study was undertaken to find out the prevalence of common micro-organisms causing UTI and to determine the antimicrobial sensitivity pattern among the patients attending Baba Raghav Das Medical College and associated Nehru Hospital, Gorakhpur, Uttar Pradesh, India.

Duration
Prospectively designed one year study duration i.e. from January 2015 to December 2015

Specimen
Clean-Catch midstream urine of the patients were collected in a sterile container and immediately transported to IJBAR (2016) 07 (06) www.ssjournals.com the bacteriology laboratory. Guidelines for proper specimen collection were given to all patients on a printed card. [7] 2.3 Inclusion criteria All patients clinically suspected of having UTI were subjected to semi-quantitative culture and sensitivity; all culture positive micro-organisms among any age group irrespective of sex distribution were included, except candiduria.

Specimen Processing:
Urine specimens were cultured semi-quantitatively for isolation of the microbial agents of UTI on blood agar and MacConkey agar media (Himedia, India & Merck, Germany). All the bacteria isolated from urine in this study were identified using conventional biochemical tests. [8]

Bacterial identification
Semi-quantitative culture of urine samples was done by calibrated loop method on 5% sheep blood agar and MacConkey agar plates and incubated in aerobic conditions at 37°C for 24-48 hours. The urine cultures of colony count >10 5 colony forming units (CFU)/mL with no more than two species of microorganisms were considered as positive for UTI and cultures showing growth of more than two types of bacteria were considered contaminated. Positive cultures were further subjected to battery of various biochemical reactions for identification up to species level. [9]

Criteria for the selection of the ESBL producing strains
The isolates were tested for their susceptibility to the third generation cephalosporins e.g. ceftazidime (30 μg), cefotaxime (30 μg) and ceftriaxone (30 μg) by using the standard disc diffusion method, as per CLSI guidelines. If a zone diameter of < 22 mm for ceftazidime, < 27 mm for cefotaxime and < 25 mm for ceftriaxone were found, the strain was considered to be "suspicious for ESBL production". Simultaneously, the ESBL status was further confirmed by disk using beta-lactam antibiotic plus betalactamase inhibitor combination by phenotypic disk diffusion test. [10]

Statistical analysis
The statistical analysis was performed by using the Chi-square test and p-value of less than 0.05 was considered as statistically significant.

Result
A total of 2217 samples were collected in the study period of one year, of which 1424 (64.23%) were from female and 793 (35.77%) were from male. Culture positivity was 43.61% (967/2217) for significant bacteriuria irrespective of all age groups and sex. Among male patients maximum number of positivity were from age group less than one year, while in female patients the maximum number of positivity were found in age group 1-30 years as shown in table 1. In this study, out of 967 total culture positive samples, 648 (67.01%) Gram-negative bacilli and 319 (32.99%) Gram-positive cocci were isolated.  The antimicrobial sensitivity of selected and recommended antimicrobial agents against Gram negative and Gram positive uropathogens are summarized in table 3 and 4 respectively.

Discussion
Early detection and selection of an appropriate effective antimicrobial agent is highly essential for effective management of patients suffering from UTIs. In our study, male children (<1 year) have higher incidence of UTI than female children of the same age group. This was in accordance with a similar study in which during the first year of life, the incidence of UTI in female child was low as compared to male. [11] In this study, it has been found that during reproductive age group (18-40 years) the prevalence of UTI in female was high as compared to male as similar with other study. [12] Female are more prone to UTI because of anatomic reasons; short and straight urethra and short distance between the ostium of the urethra and the anus contribute to easy colonization of the peri-urethral region with enteric bacteria. [13] High prevalence rate of UTI found in elderly female in this study with age group of more than 60 years, several factors importantly influence the occurrence of UTI among postmenopausal women. The reason for high prevalence of UTI include, a recurrent history of UTIs, nonsecretor status, and possibly other inherited predispositions, as well as urodynamic factors, especially incontinence, residual urine volume, and presence of a cystocele. [14] In the present study, overall prevalence is also high in female (46.48%) as compared to male (38.46%), which is consistent with other study. [15] Among Gram negative uropathogens, Escherichia coli was the most common isolated organism (35.80%) followed by Klebsiella pneumoniae (18.92%), which is similar with other studies. [15,16] Several factors are responsible for attachment of Enterobacteriaceae to the uroepithelium like, they colonize the urogenital mucosa with adhesin and pili. [17] In our study, among higher antibiotics imipenem (92%), ceftazidime-clavulanic acid (71%), piperacillin-tazobactam (68%) followed by amikacin (60%) and nitrofurantoin (56%) were found to be the most effective antibiotics against commonly isolated Gram negative uropathogens whereas flouroquinolones(13-17%) and oral cephalosporins (17%) found to be least sensitive, as observed in similar other study where, antibiotic sensitivity test performed for Escherichia coli, Klebsiella pneumoniae showed lowest sensitivity to cephalosporins (28%) and highest sensitivity to imipenem(100%). [18] Although all of these are parenteral antibiotics and is difficult to use in outdoor patients setting except nitrofurantoin, which is oral antibacterial agent, cheap and easily available in developing countries. E. coli was most frequently isolated uropathogen (35.80%) and consistently sensitive (63%) to nitrofurantoin as shown by other similar study. [19] The consistent and high-level susceptibility of E. coli to nitrofurantoin may be influenced by its narrow spectrum of activity, limited indication, narrow tissue distribution, and limited contact with bacteria outside the urinary tract. [20] Thus, nitrofurantoin has become an important oral agent in the treatment of uncomplicated urinary tract infections. [21] Among Gram positive isolates, Enterococcus species were the most common isolated organism (12.92%) followed by coagulase negative Staphylococcus (12.10%), in contrast with other study where Enterococcus species was found as the most frequent organism (15%), followed by Staphylococcus aureus (1%). [22] Our results indicated that only (32.99%) 319/967 of the UTI cases were caused by Gram-positive microorganisms. Enterococcus spp. was detected as a more resistant uropathogen than Staphylococcus aureus Moreover, Enterococcus spp. showed a high rate of resistance to norfloxacin (92%), ciprofloxacin (91%) and trimethoprimsulfamethoxazole (91%) but showed highest sensitivity to vancomycin (100%) and linezolid (78%), these findings are in agreement with other study. [22] Similarly, Staphylococcus aureus showed highest sensitivity to vancomycin and linezolid (100%) and (90%) respectively. Overall vancomycin and linezolid had strong antimicrobial activity against Gram positive isolates similar with other studies [22,23] whereas, trimethoprim-sulfamethoxazole are recommended as a first-line therapy for the management of uncomplicated UTIs, but our study revealed high rate of resistance (84%) against this antimicrobial agents. [24] Our study, demonstrated the highest frequency of ESBL production by Acinetobacter spp. (84%) whereas, Alyamani et al., reported (94%). [25] The second higher producer of ESBL was Pseudomonas aeruginosa (73.07%), in contrast Shaikh et al., reported 25.13% ESBL prevalence, which is much lower than our finding. [26] This shows a rising trends of ESBL in these days and the reason may be indiscriminate and rampant use of cephalosporins for the treatment of common infections, as well as horizontal transmission of resistant genes among hospital acquired bacterial strains. The prevalence of ESBL was 71% in Klebsiella species and 66% in E.coli, which is much higher as compared to other study; where 35% E. coli and 23.6 -41% Klebsiella pneumoniae were found to ESBL producers. [18,27] Most of these isolates were confirmed by combination of beta lactam and beta lactamase inhibitor and showed overall 69.13 % of ESBL producing strains.
In developing countries, the frequent prescriptions of antibiotics for the treatment of UTI and community level poor hygiene are the reasons for the ever-growing antimicrobial resistance in uropathogens. Urinary pathogens showed resistance to commonly used antibiotics like flouroquinolones but good sensitivity was observed with nitrofurantoin. The susceptibility and resistance patterns as observed in the defined geographical area should be considered before starting empirical treatment for UTI. As susceptibility pattern is changing with the change in use of type of antibiotics, a regular monitoring of antibiotic resistance pattern is very helpful in ensuring proper therapy IJBAR (2016) 07 (06) www.ssjournals.com for patient with urinary tract infections. We conclude that clinicians should encourage accurate bacteriological diagnosis of each symptomatic patient as far as possible and refer to the record of local microbial isolation and their antibiogram in cases of emergency or in areas where the culture facility is not available and to minimize the antimicrobial resistance. The microbiology laboratory could play an important role in record keeping of UTI isolates and their antibiogram, may succour clinicians for better and efficient management of these cases.