Essay: Etiology and susceptibility to antibiotics in positive urine culture.

Introduction:

Urinary tract infections are defined as microbial infiltration of the sterile urinary tract [1]. They are considered the most common bacterial infections associated with high morbidity across the globe, currently a serious public health problem [2, 3]. Urinary tract infections are more common in women due to anatomical and hormonal differences. In terms of etiology, Escherichia coli causes 80% of acute infections in patients without catheter, urinary stones and obstructive causes [1].

It displays an increasing resistance to broad-spectrum antibiotics (cephalosporins and quinolones) due to excessive prescription and development of co-resistance to other antibiotics by using them in parallel. This ESBL (extended-spectrum β-lactam antibiotics) producing strains cause bacterial resistance to most antibiotics, except carbapenems. A worrying phenomenon reported by studies on all continents show ESBLCARBA type (New Delhi metallo-β-lactam NDM-1) strains resistant to all carbapenems [4].

As a primary treatment for uncomplicated cystitis, IDSA (Infectious Diseases Society of America) 2010 guidelines recommended trimethoprim sulfamethoxazole if the local rate of resistance is < 20%, nitrofurantoin for five days or a single dose of fosfomycin [5]. Quinolones should not be routinely prescribed for the treatment of cystitis, being first intention empirical therapy in pyelonephritis [6].

The purpose of this study is to assess the most important aspects about etiology and susceptibility to antibiotics in positive urine culture.

Methods:

We performed a retrospective study by processing bacterial strains isolated from urine culture during 1.01.2014-31.05.2014 in the Laboratory of Infectious Diseases Clinic, Mures County Hospital. Data were collected from laboratory records and results were statistically and graphically processed (Excel programme).

Population studied: We included in the study bacterial strains isolated from urine culture in hospitalized or ambulatory patients. Bacterial strains isolated from the same patient were excluded from the study.

Urine Specimen Collection: urine samples were collected before initiating antibiotic therapy in sterile containers, from morning midstream urine emitted spontaneously or by urethral or suprapubic urinary bladder catheterization.

Urine Culture and Antibiotic Susceptibility Test: The used laboratory method was calibrated loop method, seeding with 1μl calibrated loop of blood agar culture medium, CLED. After incubation for 18-24 hours at 37 °C, bacterial colonies can be identified by classical biochemical methods. We only considered results with clinical significance > 100.000 UFC/ ml with one type of germ. Antibiotic susceptibility test – we used difuzimetric method (inoculum of 0.5 McFarland with antibiotic disks on Mueller-Hinton Agar medium). Clinical and Laboratory Standard Institute (CLSI) antimicrobial susceptibility testing standard was used. The interpretive standards provided by the CLSI are derives from a correlation between zone sizes observed in a test and MICs (minimum inhibitory concentration) of the species that can be tested by the disk diffusion method [7]. RENAR accredited SR ISO 15189: 2012 (Medical Laboratories Requirements for quality and competence).

The study received the approval of the Mures County Hospital ethics committee.

Results:

1130 urine samples were collected during the studied period, out of which 184 (12%) were positive. The study showed a net predominance of cases in women – 155 cases (84%) rather than males – 29 cases (16%). Considering the distribution by hospital departments, we detected an increased frequency of cases in Obstetrics and Gynecology – 81 cases (45%) and Clinical Infectious Diseases – 70 cases (38%).

Escherichia coli was the main etiologic agent isolated from urine culture in 119 cases (64.67%). Increased percentage of Enterococcus strains were isolated in 22 cases (11.95%) and Klebsiella pneumoniae in 17 cases (9.23%), followed by Streptococcus agalactiae in 15 cases (8.15%) and Proteus mirabilis 3 cases (1.63%). Enterobacter species, Acinetobacter, Pseudomonas, Staphylococcus strains were rarely isolated, with frequency below 1%.

Insert Table I

Depending on the susceptibility of the isolated strains to the main antibiotics used to treat urinary tract infections, Escherichia coli showed the highest resistance to ampicillin (73 cases – 61.34%), followed by aminoglycosides (gentamicin 32 cases- 27. 82%, tobramycin 26 cases- 22.41%) and trimethoprim sulfamethoxazole (31 cases- 26.96%). Excellent susceptibility was registered to meropenem (9 cases -100%) and a good sensitivity to nitrofurantoin (104 cases – 87.39%). ESBL-producing Escherichia coli strains were isolated.

Klebsiella pneumoniae strains showed absolute resistance to ampicillin (17 cases – 100%). We also found 4 cases with absolute resistance to piperacillin-sulbactam, and 4 cases to ticarcilin – clavulanate. Increased resistance was recorded to ampicillin – sulbactam (9 cases – 52.94%) and gentamicin (8 cases – 47.05%). In our study 5 strains of Klebsiella pneumoniae were ESBL-producing and 4 CRE (Carbapenem-resistant Enterobacteriaceae) strains were detected. We observed good susceptibility to amoxicillin- clavulanate (8 cases- 80%), cephalosporins. One strain of Klebsiella was isolated which proved resistant to all tested antibiotics, except for trimethoprim- sulfamethoxazole.

Regarding Proteus mirabilis strains isolated from urine culture, we noted maximum resistance to ampicillin and a high resistance to nitrofurantoin. The strain recorded high susceptibility to all tested cephalosporins, ampicillin-sulbactam and amoxicillin-clavulanate.

Insert Table II

Enterococcus spp. strains showed different degrees of resistance to the tested antibiotics, the highest level displayed for trimethoprim-sulfamethoxazole (21 cases – 100%), increased resistance for streptomycin (12 cases – 80%) and imipenem (6 cases – 30%). There have been 2 cases of Vancomycin-Resistant Enterococcus (VRE). We recorded increased susceptibility to last generation antibiotics (vancomycin 19 cases – 90.47%, linezolid 19 cases- 90.47%, teicoplanin 18 cases – 85.71%). Excellent susceptibility to tigecycline was depicted (20 cases – 95.23%).

Insert Table III

There were isolated 15 cases of Streptococcus (agalactiae) beta haemolitic susceptible to beta-lactam antibiotics. We also found one strain of Staphylococcus aureus susceptible to all tested antibiotics except for benzylpenicillin and one strain of Pseudomonas spp. susceptible to all tested antibiotics.

Discussion:

Community urinary tract infections have a prevalence of 0.7% in the world, their incidence being dependent on many factors including age and patient characteristics, the associated pathologies and the personal history of urinary tract infections [4]. Nosocomial urinary infections are classified in the top three places as frequent in the world [8].  Specialty studies highlight a continuous increase in urinary tract infections with resistant germs ….

Urinary tract infections, especially those uncomplicated prevail in women. Shorter urethra which facilitates colonization with gastrointestinal pathogens, sexual activity, contraceptive use and low levels of estrogen can be contributing factors [1, 3, 9, 10].

In our study the most frequent positive urocultures were recorded at the Obstetrics Gynecology Clinic. Due to anatomical and hormonal changes (ureteral dilatation, uretero-bladder reflux, urinary stasis), about 8% of pregnant women develop urinary tract infections as complication of pregnancy, with a peak in the second trimester [11, 12, 13].

Urinary tract infections can be caused by Gram-pozitive germs, fungi, but studies show that most frequently are caused by Gram-negative bacteria. They associate many factors responsible for attachment to urinary epithelium, able to colonize the urogenital mucosa with adhesins, pili, fimbriae and P-1 receptor phenotype in the blood [14]. Studies highlight Escherichia coli the most commonly isolated pathogen in both previously healthy patients and those with associated pathologies [15]. The main pathogen agent, isolated in over 80% of community-acquired urinary tract infections, is Escherichia coli. Pathogens commonly associated with uncomplicated urinary tract infection are species of Klebsiella, Proteus mirabilis, Enterococcus faecalis, group B Streptococcus, Staphylococcus saprophyticus [16, 17].

Hospital-acquired infections are related to the presence of urethral catheter in 70-80% of cases [18, 19]. Echerichia coli is also the most common isolated pathogen agent followed by the frequency of Enterococcus spp., Klebsiella pneumoniae, Staphylococcus aureus, Proteus mirabilis, Pseudomonas aeruginosa.[17] Proteus mirabilis is common in chronically catheterized patients. It was isolated in 40% of patients and shows the long-term trend of persistent infection [19].

The present study highlights the susceptibility of Echerichia coli strains to amoxicillin-clavulanate, cephalosporins, quinolones, trimethoprim-sulfamethoxazole, which allows their use in uncomplicated urinary tract infections. There are geographical areas where Echerichia coli register increased resistance to both penicillin and cephalosporins [20]. The resistance of Echerichia coli strains to quinolones is extremely widespread in the world [8], a worrying phenomenon by lowering the therapeutic effectiveness of this class of antibiotics considered by choice in the treatment of urinary tract infections. Uncontrolled consumption and empiric prescription of antibiotics, as well as lack of patient adherence and antibiotic quality lead to the appearance of ESBL-producing strains [21]. Good susceptibility to nitrofurantoin, as confirmed by other similar studies [4, 22] can be explained by the fact that this substance was not used routinely for the treatment of urinary tract infections over a long period of time. The highest percentage of resistance was proved in the case of ampicillin, probably due to more frequent use of antibiotics in recurrent infections; patients are more exposed to self-medication and repeated courses of antibiotic therapy in hospital.

In our study, resistance profile of Klebsiella pneumoniae strains showed high resistance to the tested antibiotics. With the introduction of carbapenems, antibiotic resistance in the case of Klebsiella pneumonia strains began to spread. This is a particularly worrying phenomenon because carbapenems are last-line antibiotics and treatment options for patients infected with carbapenem-resistant bacteria are severely limited [23, 24, 25].

Strains of Enterococcus species register different degrees of resistance to antibiotics, ei fiind obisnuit rezistenti la antibiotice cum ar fi penicilinele, cefalosporinele, trimethoprim [17, 26]. In recent years, Enterococcus species have developed glycopeptide resistance, the incidence of Vancomycin-Resistant Enterococcus species has almost been doubled, with 30% of the cases reported as vancomycin resistant [27].

In the case of Proteus mirabilis strains isolated from urine samples, there is a tendency of increase in the resistance to certain classes of antibiotics (beta lactams, quinolones, aminoglycosides, nitrofurantoin, trimethoprim-sulfamethoxazole) due to a bacterial embedding mechanism to urinary calculi and adherence to catheter. Current recommendations in acute uncomplicated cystitis, whose agent pathogen is Proteus mirabilis, indicates treatment with trimethoprim sulfamethoxazole for three days if resistance rate is less than 10-20%. When resistance rate exceeds 20%, nitrofurantoin, fosfomycin and quinolones are recommended as alternative treatment options. In case of complicated urinary tract infections associated with catheters, there are no specific treatment recommendations [28].

The study has several limitations: studied population is not homogeneous due to laboratory servicing for certain clinics. A greatly increased incidence in women has been noticed, possibly also connected to the bacterial urinary determinations in Clinic of Obstetrics and Gynecology. Samples from Intensive Care Unit were tested in another laboratory so we do not have records of a higher percentage of multidrug resistant strains. Since data were collected from laboratory records, we could not fully evaluate patients characteristics.

Conclusions:

Etiologic spectrum of urinary tract infections is dominated by Escherichia coli and Enterococcus species, followed by Klebsiella pneumoniae. Positive urine cultures prevailed in women and were more common in the Clinic of Obstetrics and Gynecology.

Echerichia coli strains isolated from urine culture display high ampicillin resistance, allowing the recommendation to avoid its use as first-line regimen. Klebsiella strains isolated from urine culture displayed resistance to ampicillin in all cases.

Bacterial strains most often isolated from urine culture retain their susceptibility to nitrofurantoin, making it advisable for the treatment of mild urinary tract infections; susceptibility to carbapenems recommends these antibiotics in severe urinary tract infections, caused by multi-drug-resistant bacteria.

Regarding antibiotic susceptibility of isolates from urine cultures, we registered great susceptibility to last generation antibiotics, which maintain the quality of drugs of last resort.

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