Nitrofurantoin susceptibility profile versus other antibiotics tested in uropathogens-a retrospective study from India .

ABSTRACT


Introduction
Urinary Tract Infection (UTI) is one of the most common bacterial infections encountered by clinicians worldwide.The emergence of multidrug-resistant uropathogens necessitates a review of their susceptibility profiles.This study aims to assess the susceptibility trends of uropathogens to a panel of drugs, with special emphasis on Nitrofurantoin (NFT).

Methods
A retrospective analysis was conducted on 2,099 mid-stream clean catch urine samples processed by standard microbiological methods.Clinical and Laboratory Standards Institute (CLSI) guidelines (2019) were followed.Statistical analysis was performed.

INTRODUCTION
Urinary Tract Infection (UTI) is one of the most common bacterial infections encountered by clinicians worldwide, significantly contributing to the workload in clinical microbiology laboratories (1,2).UTI is a general term referring to infection or inflammation of any part of the urinary tract (1).It is divided into two types based on the site of involvement: upper UTI and lower UTI .
The prevalence of UTI varies depending on age, sex, catheterization, hospitalization, and previous exposure to antimicrobials (4 The overall prevalence of UTI ranges from 8.7% to 90.1% (5).UTI is more common in females than in males due to factors such as sexual activity, hormonal changes, a shorter urethra, and the proximity of the urethral orifice to the anus (3,6).
UTIs are most often caused by bacteria but can also involve fungal, viral, and parasitic pathogens.Gram-negative bacteria cause 90% of UTI cases, while Gram-positive bacteria account for only 10%.Escherichia coli (70-95%) is noted as the predominant uropathogen (7).
The increase in multidrug-resistant (MDR) organisms has become an alarming situation globally, including in India.A study from South India found an overall prevalence of MDR organisms at 54% (8).Another study from India highlighted that 23% of uropathogenic E. coli were ESBL producers, and around 12% were MDR uropathogenic E. coli (9).A study from eastern North India reported that 96% of the isolated uropathogens were MDR organisms, with 40.4% of E. coli isolates being ESBL producers (10).Commonly used oral antibiotics like cotrimoxazole and fluoroquinolones are no longer effective, especially in inpatient settings (11).Moreover, higher-end antibiotics such as third-generation cephalosporins and carbapenems are losing effectiveness due to the rapid emergence of Extended Spectrum Beta-Lactamases (ESBLs) and carbapenemases, respectively (11,12).Furthermore, the development of novel antibiotics is lagging.This situation has necessitated revisiting age-old antimicrobials such as nitrofurantoin for their activity against these multidrug-resistant uropathogens (3,13).
Nitrofurantoin is an example of an oral antibiotic that was approved by the Food and Drug Administration (FDA) in 1953 for the treatment of lower UTIs.It was widely used for uncomplicated UTIs until the 1970s when trimethoprim-sulfamethoxazole and newer betalactam antibiotics became available (14).Due to the paucity of data regarding effective drugs for MDR uropathogens, this study was undertaken to assess the susceptibility trends of all uropathogens isolated to a panel of drugs, with special reference to nitrofurantoin.

MATERIALS AND METHODS
This retrospective study was conducted in the Department of Microbiology at a tertiary care hospital in New Delhi, India.A convenient sampling method was used, including 2,099 urine samples received over six months from December 2019 to May 2020.Culture sensitivity testing was performed as part of routine diagnostics during patients' inpatient stays, for which patient consent was not required.All data from test requisition forms were recorded in laboratory registers and maintained in desktop records, from which it was analyzed.

Ethical approval and consent
Since this was a retrospective analysis, departmental-level approval was obtained from the bacteriology section.According to institutional policy, departmental ethical clearance suffices for retrospective studies; hence, both departmental and sectional clearances were obtained.Every effort was made to ensure patient confidentiality during data collection.All available measures were taken to maintain confidentiality related to patient details while compiling data for the study.
The study included urine samples from adult inpatients (aged >18 years) of both sexes.Urine samples from outpatient attendees and individuals under 18 years were excluded from the study.

Specimen collection and processing
Midstream urine samples were collected in wide-mouth, leak-proof, screw-capped sterile universal containers with properly filled test requisition forms, duly signed by the referring doctor (15).Samples were processed within 2-4 hours of collection for aerobic culture and sensitivity testing.Uropathogens were isolated on Blood agar and MacConkey agar using a 0.01 mm calibrated loop for the semi-quantitative method.The plates were incubated overnight at 37°C and observed for discrete growth for characterization and identification of the pathogen.Colony counts >10^5 colony-forming units (CFU)/ml in patients with no risk factors were considered significant, while in symptomatic patients (where history was provided in test requisition forms), colony counts of >10^3 CFU/ml were considered significant (15).Bacterial identification was performed using colony characteristics, Gram staining, catalase, oxidase, coagulase production, and various biochemical tests, including Triple Sugar Iron, indole, citrate utilization, urea hydrolysis, sulfide-indole motility (SIM) medium, mannitol salt agar, DNase, and bile esculin hydrolysis (16).

Antibiotic susceptibility testing
Antibiotic susceptibility testing was performed on each isolated bacterium using the Kirby-Bauer disc diffusion method according to the Clinical and Laboratory Standards Institute

Quality control and Quality assurance
Standard Operating Procedures (SOPs) were strictly followed, ensuring that culture media and antibiotic disks met expiration date and quality control parameters.Quality control was performed according to SOPs.Each new lot was checked with the reference strains: E. coli (ATCC 25922), S. aureus (ATCC 25923), and P. aeruginosa (ATCC 27853) (17).

Statistical analysis
SPSS software version 21.0 was used for data analysis.A two-tailed Chi-square test was used to compare categorical variables.A p-value <0.05 was considered significant.

DISCUSSION
This study provides important information regarding antimicrobial susceptibility trends in both Gram-negative and Gram-positive uropathogens.Our findings may serve as a foundation for exploring the administration of Nitrofurantoin (NFT) among multidrugresistant uropathogens.
A predominance of female patients was observed, similar to studies conducted by Akhter et al. and Naik et al. (19,20).Since the majority of samples were from females, the culturepositive cases were also higher in females, at 9.3%, consistent with the findings of Akhter et al. (19).The high prevalence of UTI in females of reproductive age is attributed to the proximity of the urethral meatus to the anus, a shorter urethra, sexual intercourse, incontinence, and poor toilet hygiene (19,20).
The most commonly isolated Gram-positive uropathogen was Enterococcus spp. at 15.1%, comparable to results seen by Akhtar et al. (19).We reported that out of the total VRE isolates, 40% were sensitive to Nitrofurantoin (2 out of 5 isolates).The incidence of VRE in the current study is 2.34%, comparable to that obtained by Meena et al. (2.7%) (13).
Due to the retrospective nature of the study, crucial data and result findings cannot be correlated with the clinical diagnosis, associated signs, symptoms, and comorbidities.However, clinicians have been informed about the summary findings of the drugs, which may be beneficial in future probable cases of UTI.
a only for Staphylococcus spp.bonly for Enterococcus spp.

Table 3 .
Susceptibility profile of uropathogens to nitrofurantoin.
(3),24)9 samples received, 212 (10.22,23)re culture-positive, which aligns with other studies by Kasew et al.,Nahar et al., and Tesfa et al. (21,22,23).Differences in prevalence could be due to varied geographical distribution, length of study, sample size, and seasonal variation(23).The study reinforces the well-known fact that E. coli is the leading uropathogen, comparable to other studies by Derbie et al. and Kasew et al.(21,24).We observed high resistance among Gram-negative uropathogens towards ciprofloxacin, ceftriaxone, and cotrimoxazole, which are commonly used antibiotics for UTI treatment.These results were comparable to those of Goyal et al.(3).Gram-negative uropathogens remained predominantly sensitive to Netilmicin, Imipenem, and Amikacin, which are parenterally administered and reserved for inpatient use.Among Gram-negative uropathogens, NFT, an age-old oral drug, showed good sensitivity, similar to the results seen by Goyal et al.(3).All Gram-positive isolates were sensitive to Linezolid, whereas Vancomycin and Nitrofurantoin each showed 92.45% sensitivity.Goyal et al. (3) found contrasting results with maximum sensitivity to Nitrofurantoin (76%), followed by Linezolid (69%) and Vancomycin (58%).Additionally, we highlighted that Staphylococcus aureus and CoNS were 100% sensitive to Nitrofurantoin, while Enterococcus spp.showed 87.5% sensitivity.The majority of E. coli isolates were sensitive to Nitrofurantoin at 90.5%, which is consistent with findings by Goyal et al.,Gautam et al., and Neelima A et al.