Key findings:

The key findings of this study on urinary tract infections (UTIs) include: Klebsiella pneumoniae as a common causative agent, particularly among non-E. coli UTIs; high resistance to antibiotics such as Ampicillin, Ceftazidime, and Azithromycin; and sensitivity to Amikacin, Gentamicin, and Imipenem, highlighting the need for effective treatment strategies to combat antimicrobial resistance.

What is known and what is new?

The known aspect in this abstract is the prevalence of urinary tract infections (UTIs) caused by pathogens like Escherichia coli and Klebsiella pneumoniae. The new contribution is the study's focus on antimicrobial resistance patterns of Klebsiella pneumoniae in UTI patients, highlighting the emergence of resistance to common antibiotics and the need for effective treatment strategies.

What is the implication, and what should change now?

The implication of this study is the concerning antimicrobial resistance of Klebsiella pneumoniae in UTI patients, necessitating the development of tailored treatment protocols to address resistance patterns. Changes needed include implementing antimicrobial stewardship programs, promoting judicious antibiotic use, and exploring alternative treatment options to combat the rising challenge of antibiotic resistance in UTIs.

Urinary tract infections (UTIs) are one of the most common bacterial infections, affecting millions of individuals worldwide. Among the various pathogens associated with UTIs, Klebsiella pneumoniae has emerged as a significant cause of concern due to its increasing prevalence and alarming rates of antibiotic resistance. K. pneumoniae is a Gram-negative bacterium that can colonize the urinary tract, leading to a range of infections, from uncomplicated cystitis to severe pyelonephritis [1]. Klebsiella pneumonia, belonging to the family Enterobacteriaceae, is a natural inhabitant of the gastrointestinal tract microbiome of healthy humans and animals [2].  K. pneumoniae prevalence in UTIs has been rising globally, posing a substantial burden on healthcare systems and highlighting the need for effective management strategies. Furthermore, the emergence of multidrug-resistant strains of K. pneumoniae has complicated the treatment of UTIs, making it imperative to understand the antibiotic resistance patterns of this pathogen [3]. Several studies have indicated a growing trend of antibiotic resistance among K. pneumoniae isolates from UTIs, with resistance to commonly prescribed antibiotics such as fluoroquinolones, cephalosporins, and carbapenems. This resistance not only limits treatment options but also increases the risk of treatment failure and recurrent infections [4]. K. pneumoniae has a variety of antibiotic resistance mechanisms and is a common pathogen causing hospital-acquired surgical wound infections, digestive tract infections, and community-onset infections, which can cause outbreaks of nosocomial infection [5]. The global drug resistance rate of K. pneumonia has reached as high as 70%, and the infection-related fatality rate has also reached 40%~70% [6]. Recently, multiple-drug resistance (MDR) K. pneumoniae and carbapenem-resistant K. pneumoniae (CRKP) have emerged as a major global public health problem [7]. The current study is designed to explore the infection of K. pneumonia and the antimicrobial resistance of UTI patients. A total of 77 urine specimens were collected from patients (one specimen/ patient) attending Al-Karama Teaching Hospital in Al-Kut, Wasit Province, clinically diagnosed as having UTI. Of these patients, 43 were females and 34 were males (aged from 1 month to 80 years), who were infected with Klebsiella pneumonia during the period from 10th January to 27th February 2024.

2.1 Samples' collection

A total of 77 urine specimens were collected from patients attending Al-Karama Teaching Hospital in Al-Kut, Wasit Province, clinically diagnosed as having UTI. Of these patients, 43 were females and 34 were males (aged from 1 month to 80 years), during the period from 10th January to 27th February 2024.All urine samples submitted for culture were processed in our laboratory using a fully automated plating and streaking instrument, on nutrient agar and blood agar of a midstream, 10 mL urine aliquot for cultivation. At 37°C, plates were incubated aerobically for 24–48 hours.

2.2 Culture Blood agar nutrient agar 

Urine culture is the gold standard for diagnosing UTI. Different versions of the calibrated loop/surface streak method have been used since the 1960s to semi-quantify, isolate and start a presumptive identification of the microorganisms present in a urine specimen. It is important to iterate that all samples should first be subject to a dipstick testing and/or microscopic examination to look for the presence of nitrites, white blood cells, red blood cells or bacteria.

2.3 Culturing Method

1. Tipping over the container to re-mixing the urine sample was done.
2. Removing the cap and dipping the end of a sterile 1-µL inoculating loop (white) into the urine and removing it vertically making sure that there is no urine up the loop were achieved.
3. The tipping and spreading the inoculum over the surface of a standard nutrient agar plate (60 × 15 mm) prepared according to the instructions of the manufacturing company was done. A single streak across the center was made. Then, spreading the inoculum evenly distributed in a diagnostics cross-zigzag arrangement to the primary streak was fulfilled.
4. Re-dipping the end of the same 1-µL loop into the urine and removing it vertically making sure that there is no urine up the loop was carried out.
5. Tipping and spreading the inoculum over the surface of a glucose-topped Blood agar plate (60 × 15 mm) was done for each sample. 
6. Re-dipping the end of the same 1-µL loop into the urine and removing it vertically making sure that there is no urine up the loop was done.
7. Tipping and spreading the inoculum over the surface of a standard Blood agar plate (60 × 15 mm) prepared according to the instructions of the manufacturing company was done. 
8. Incubation the plates aerobically at 35–37 ◦C for at 18-24 h was achieved.
9. In the following day, the number of colonies on the surface of each medium was counted. Each colony growing on the agar plate represents one colony forming unit (cfu)/µL (according to the size of the loop), which is equal to 1000 cfu/mL.
10. The nutrient agar is the primary medium used for counting colonies that show sensitivity for multiple disks of antibiotics

3.1 Study Group Characteristic

All urine samples submitted for culture underwent thorough processing in the laboratory, and upon incubation overnight, the resulting bacterial colonies were observed and documented. The presence of bacterial growth in the cultures indicated potential urinary tract infections or other urinary system-related conditions. The identification and characterization of these bacterial colonies are crucial steps in determining the causative agents of infection and guiding appropriate treatment strategies. Each colony's morphology, size, color, and other characteristics were carefully examined and recorded to facilitate accurate identification. The following figure 1 shows Klebsiella colonies.

Fig. 1: Klebsiella growth of a urine sample on blood agar after 24 hours of incubation

Selected colonies of Klebsiella were subcultured onto Muller Hinton agar plates to conduct antibiotic susceptibility testing and establish resistance patterns. This step is essential in determining the most effective treatment options for infections caused by Klebsiella bacteria. By exposing the isolated colonies to various antibiotics commonly used in clinical practice, healthcare professionals can assess the bacteria's sensitivity or resistance to these drugs. The results of these tests are illustrated in figure (2).

Fig. 2: Inhibition zones of random antibiotics against the bacterial growth of urine sample after incubation overnight

The final study group consisted of 77 patients diagnosed with a UTI. There were 43 females (55.84%) and 34 males (44.15%). Patients’ age varied from 1 month to 94 years. In the whole study group, there were 34 age 0-18 year (44.15%) , 16 person age 19-40 year ( 20.77%) , 14 person age 41-60 year ( 18.18%) , and 7 over 60 year ( 9.1%). Figure (3).

Fig. 3: Distribution of bacterial infections among age groups.

Urinary tract infections (UTIs) are among the most frequent community-acquired infections worldwide. E.coli is the most common UTI pathogen followed by Klebsiella although underlying host factors such as patients' age and gender may influence prevalence of causative agents. In this study, 77 consecutive urine samples were received over a two-month period. Figure 4

Fig. 4: The correlation between UTI infection, Age and Sex .

Data stratification according to both age and gender showed isolation rates to be lower in both males aged ≥60 years. In conclusion, both patients' age and gender are significant factors in determining UTIs etiology. Furthermore, the correlation between UTI and (age and sex) can be supported by another study that focused on only one microorganism E. coli [8]. 

3.2 Etiology of a UTI

The most common causative agent was E. coli, responsible for 31.16% of UTIs in the whole study group. Among non-E. coli UTIs, the dominating causative bacteria were Klebsiella spp. (22.07%), followed by Staph spp. Candida spp. Proteus spp. and Strep spp. The prevalence of uropathogens identified in all analyzed cultures is presented in figure (5).

Fig. 5: Prevalence (%) of bacterial pathogens causing a UTI in the study group.

All urine samples submitted for culture were processed in our laboratory using a fully automated plating and streaking instrument, the Copan WASP. The WASP processor used 10 L sample loops on blood agar and nutrient agar plates to plate the first 10 mL urine aliquot for cultivation. At 37°C, plates were incubated aerobically for 24–48 hours. Leukocyte esterase test results that were positive and had a UTI pathogen growth >104/mL were used to classify urine samples as positive. It was also deemed positive when two different bacterial species were recovered with similar growth rates, >104/mL, and a positive leukocyte esterase test result. The species with a predominant growth was considered clinically significant when the bacterial load of one species was >104/mL and lower for the other. Samples that contained more than two microorganisms, however, were regarded as contaminated. On the VITEK 2 Compact system, the AST was carried out. 

3.4. Antibiotic Resistance Patterns for Klebsiella spp.

Klebsiella spp. was the second most commonly isolated species among Gram-negative bacteria (22.07%). A clinical isolate of klebsiella spp. was resistant to Ampicillin (58%) , Ceftazidime (35%) and Azithromycin (17%) , and was sensitive to Amikacin (47%) , Gentamicin (23.5%) and Imipenem (23.5%) as shown in the figure (6)

Fig. 6: Resistant and sensitive Klebsiella to different antibiotic

By the figure shown above we can see that the most patient with Klebsiella infection are sensitive to Amikacin, these results are matching the study [9] on the clinical and microbiological efficacy of amikacin treatment in adult patients with lower UTIs due to ESBL-producing Escherichia coli or Klebsiella pneumonia.  The clinicians should keep in mind that amikacin treatment is an efficient and safe alternative treatment option before the carbapenem treatment especially in patients with lower UTIs caused by ESBL-producing Escherichia coli or Klebsiella pneumonia that are resistant to all oral antibiotics.

UTIs are a common problem in clinical practice. Involvement of renal parenchyma may lead to inflammation and renal scarring, with long-term consequences such as hypertension and impaired renal function [10,11,12]. Thus, proper management of a UTI includes promptly starting empiric antibiotic therapy, which is essential to treat acute infection and prevent its possible complications [13].The most common causative agents for a UTI in our study were E. coli and other enteric bacteria, such as Klebsiella spp.,  Enterococcus spp., and Enterobacter spp., which is in line with previous observations [14].  When choosing the optimal therapy for UTI, local recommendations for antibiotic therapy and individual assessment of each patient’s risk factors for non-E. coli and drug-resistant bacteria infection should be carefully evaluated [15]. This matches our study as it found that E.coli is the most common UTI pathogen followed by Klebsiella although underlying host factors such as patients' age and gender may influence prevalence of causative agents. 

In different European regions, the percentage of E. coli strains isolated from patients with a UTI resistant to amoxicillin/clavulanic acid varied from 12.2% in Greece [16] to approximately 25% in Italy [17] and Spain [18]. Hrbacek et al. (2020) reported resistance of E. coli to amoxicillin/clavulanic acid of 10% in Central Europe in urine cultures of adult patients hospitalized in the urology department [19]. The alarming trend of increasing antibiotic resistance of E. coli in patients diagnosed with a UTI within a relatively short period and with a velocity of approximately 2% per year was reported by Dejonckeheere et al. [20].

Several recently published studies describe the alarming trend of decreasing sensitivity to antibiotics.  Although the sensitivity to third- and fourth-generation cephalosporins remains relatively high [17], some studies revealed increasing resistance with an estimated rate of 1% per year [20]. This trend may have a significant implication for clinical decisions in the future. Changes in antimicrobial resistance patterns in a relatively short period suggest the need to frequently update local recommendations for empiric antibiotic therapy in UTIs.

According to a study on resistance trends of Klebsiella pneumoniae causing Urinary Tract Infections in Chongqing, 2011–2019 [21], conducted that the rate of extended-spectrum β-lactamase (ESBL)-producing K. pneumoniae fell from 48.4% in 2011 to 32.9% in 2019, and a marked jump of resistance was seen in carbapenems from 2.2% to 18.0%. The peak of carbapenem resistance rate (22.6%) to K. pneumoniae was observed in 2017 along with a low ESBL-producing rate (30.9%). Piperacillin/tazobactam and cefepime resistance levels went up from 4.4% to 25.7% and from 18.2% to 30.5%, respectively. Moreover, the K. pneumoniae isolates resistance rate to carbapenems and amikacin gradually grew up, showing their peaks in 2017, and then dropped year by year. However, ceftazidime and aztreonam resistance levels were relatively stable, fluctuating between 21.8% and 35.6%, 32.2% and 39.4%, respectively). 

These findings are matching our study where it shows Klebsiella spp. was the second most commonly isolated species among Gram-negative bacteria (22.07%). A clinical isolate of klebsiella spp. was resistant to Ampicillin (58%), Ceftazidime (35%) and Azithromycin (17%), and was sensitive to Amikacin (47%), Gentamicin (23.5%) and Imipenem (23.5%) respectively. 

The risk of prescribing empirical antibiotics to which the cultured pathogen was found resistant at an antibiogram was higher in cases of a UTI caused by Klebsiella spp., Enterobacter spp., and mixed organisms [22]. 

The clinical outcome of inadequate antibiotic therapy differed between antibiotic groups. Among the most frequently chosen initial antibiotic therapy we recommend is Amikacin. Based on our findings, we would suggest avoiding Ampicillin as a treatment. the dose of Amikacin should be carefully adjusted.

Another problem in the management of UTIs is the prevention of recurrent infections in particular groups of patients. There is no consensus regarding the prescription of antibiotic prophylaxis. Recent studies search for effective non-antibiotic prophylaxis methods, including dietary supplements, probiotics, immuno-stimulants, and vaccines [23]. However, more research is required to draw evidence-based recommendations for preventing recurrent UTIs in clinical practice.

In conclusion, the prevalence of Klebsiella pneumoniae in urinary tract infections (UTIs) is a significant concern due to its association with antibiotic resistance patterns. The emergence of multidrug-resistant strains of K. pneumoniae poses challenges for healthcare providers in treating UTIs effectively and highlights the importance of antimicrobial stewardship practices.

Our study concluded that Klebsiella spp. infection is the second most commonly isolated species among Gram-negative bacteria. The clinical isolate of Klebsiella spp. exhibited significant resistance to certain antibiotics, notably Ampicillin and Ceftazidime, while showing sensitivity to others such as Amikacin, Gentamicin, and Imipenem. These results emphasize on the critical role of careful antibiotic management and continuous monitoring to guide effective treatment strategies against multidrug-resistant pathogens like Klebsiella spp.Collaboration between healthcare providers, microbiologists, epidemiologists, and policymakers is essential to combating the spread of resistant strains and preserving the effectiveness of antibiotics for the treatment of UTIs caused by K. pneumoniae.

Funding: No funding sources.

Conflict of interest: None declared.

Ethical approval: The study was approved by the Institutional Ethics Committee of Kut University College.

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