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Bacterial and fungal characteristics of urinary tract infections in pediatric patients

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Adane Bitew, 1 Nuhamen Zena, 2 Abera Abdeta31 Department of Medical Laboratory Sciences, Faculty of Health Sciences, Addis Ababa University, Addis Ababa, Ethiopia; 2 Microbiology, Millennium School of Medicine, St Paul’s Hospital, Addis Ababa, Ethiopia Department; 3 National Reference Laboratory for Clinical Bacteriology and Mycology, Ethiopian Institute of Public Health, Addis Ababa, Ethiopia Corresponding Author: Abera Abdeta, National Reference Laboratory for Clinical Bacteriology and Mycology, Ethiopian Institute of Public Health, PO Box: 1242, Addis Ababa, Ethiopia, +251911566420, email [email protected] Background: UTIs are common infections in paediatrics.Knowledge of common causes of urinary tract infections, their patterns of antimicrobial susceptibility, and associated risk factors in specific settings can provide evidence for appropriate treatment of cases.Objectives: This study aimed to determine the common etiology and prevalence of associated uropathogens and urinary tract infections, as well as the antibiotic susceptibility profiles of bacterial isolates, and to identify risk factors associated with urinary tract infections in pediatric patients.Materials and methods: The study was conducted from October 2019 to July 2020 at the Millennium School of Medicine, St. Paul’s Hospital.Patient urine is collected aseptically, inoculated onto media, and incubated at 37°C for 18-48 hours.Bacteria and yeast were identified according to standard procedures.Antibiotic susceptibility testing of bacterial pathogens using the Kirby Bauer disc diffusion method.Descriptive statistics and logistic regression were used to estimate raw ratios with 95% confidence intervals.P-value results: Significant bacterial/fungal growth was observed in 65 samples with a prevalence of 28.6%, of which 75.4% (49/65) and 24.6% (16/65) were bacterial and fungal pathogens, respectively.About 79.6% of the bacterial etiologies were Escherichia coli and Klebsiella pneumoniae.Resistance was highest to ampicillin (100%), cefazolin (92.1%) and trimethoprim-sulfamethoxazole (84.1%), which are commonly used empirically in Ethiopia.Length of hospital stay (P=0.01) and catheterization (P=0.04) were statistically associated with urinary tract infection.Conclusions: Our study observed a high prevalence of urinary tract infections.Enterobacteriaceae are the leading cause of urinary tract infections.Length of hospital stay and catheterization were significantly associated with urinary tract infection.Both Gram-negative and Gram-positive bacteria were extremely resistant to ampicillin and trimethoprim-sulfamethoxazole.Keywords: Antibiotic susceptibility patterns, Pediatrics, Urinary tract infections, Ethiopia
Urinary tract infections (UTIs) caused by bacteria and yeast are one of the most common urinary tract diseases in children.In developing countries, it is the third most common infection in the pediatric age group after respiratory and gastrointestinal infections.2 Intestinal infections in children are associated with short-term morbidity, including fever, dysuria, urgency, and low back pain.It can also lead to long-term kidney damage, such as permanent kidney scarring and long-term problems, including high blood pressure and kidney failure.3 Wennerstrom et al15 described renal scarring in approximately 15% of children after a first UTI, underscoring the importance of prompt diagnosis and early treatment of urinary tract infections.In addition, the expenditure on urinary tract management associated with healthcare is quite high.3,4 Numerous studies of pediatric UTIs in different developing countries have shown that the prevalence of UTIs varies from 16% to 34%.5-9 In addition, up to 8% of children aged 1 month to 11 years will develop at least one UTI10, and up to 30% of infants and children are known to have recurrent infections within the first 6-12 months after the initial UTI .11
Gram-negative and Gram-positive bacteria, as well as certain Candida species, can cause urinary tract infections.E. coli is the most common cause of urinary tract infections, followed by Klebsiella pneumoniae.12 Studies have shown that Candida species, especially Candida albicans, remain the most common cause of Candida UTIs in children.13 Age, circumcision status, and indwelling catheters are risk factors for UTIs in children.Boys are more vulnerable in the first year of life, after which, due to differences in sex organs, the incidence is mainly higher in girls, and uncircumcised male infants are at higher risk.1,33 Antibiotic susceptibility patterns of uropathogens vary over time, patient geographic location, demographics, and clinical characteristics.​​​1
Infectious diseases such as UTIs are thought to be responsible for 26% of global deaths, 98% of which occur in low-income countries.14 A study of pediatric patients in Nepal and India reported an overall prevalence of UTIs of 57%15 and 48%,16.A hospital study of South African children showed that urinary tract infections accounted for 11% of healthcare infections.17 Another study in Kenya found that urinary tract infections accounted for approximately 11.9% of the burden of febrile infections in young children.18
Few studies have identified UTIs in paediatric patients in Ethiopia: studies at Hawassa Referral Hospital, Yekatit 12 Hospital, Felege-Hiwot Specialist Hospital and Gondar University Hospital showed 27.5%, 19 15.9%, 20 16.7%, 21 and 26.45% and 22, respectively.In developing countries, including Ethiopia, the lack of urine cultures at various levels of sanitation remains impractical because they are resource-intensive.Therefore, the pathogen spectrum of UTI and its drug susceptibility profile in Ethiopia are hardly known.To this end, this study aimed to determine the prevalence of urinary tract infections, analyze bacterial and fungal pathogens associated with UTIs, determine the antimicrobial susceptibility profiles of bacterial isolates, and identify major susceptibility factors associated with UTIs.
From October 2019 to July 2020, a hospital-based cross-sectional study was conducted at the Paediatrics Department of St Paul’s Hospital Millennium Medical College (SPHMMC), Addis Ababa, Ethiopia.
During the study period, all pediatric inpatients and outpatients were seen in pediatrics.
During the study period, all pediatric inpatients and outpatients with UTI signs and symptoms attended the study site.
Sample size was determined using a single-proportion sample size calculation formula with a 95% confidence interval, 5% margin of error, and the prevalence of UTIs in earlier work [15.9% or P=0.159)] Merga Duffa et al20 in Addis Ababa, as shown below.
Z α/2 = 95% confidence interval critical value for normal distribution, equal to 1.96 (Z value at α = 0.05);
D = margin of error, equal to 5%, α = is the level of error people are willing to tolerate; plug these into the formula, n= (1.96)2 0.159 (1–0.159)/(0.05)2=206 and assume 10% unanswered where n = 206+206/10 = 227.
A convenient sampling method was used in this study.Collect data until the desired sample size is achieved.
Data were collected after obtaining written informed consent from parents.The sociodemographic characteristics (age, gender, and place of residence) and associated risk factors (catheter, previous UTI, human immunodeficiency virus (HIV) status, circumcision, and length of hospital stay) of study participants were collected by qualified nurses using pre-specified data. A structured questionnaire for the test.Signs and symptoms of the patient and underlying disease were recorded by the attending pediatrician.
Before analysis: sociodemographic characteristics (age, gender, etc.) and clinical and treatment information of study participants were collected from questionnaires.
Analysis: The performance of the autoclave, incubator, reagents, microscope, and microbiological quality of the medium (sterility of the medium and growth performance of each medium) were assessed according to standard procedures prior to use.Collection and transport of clinical samples are performed after aseptic procedures.The inoculation of clinical samples was performed under a secondary safety cabinet.
Post-Analysis: All extracted information (such as laboratory results) is checked for eligibility, completeness and consistency and recorded before entering statistical tools.Data is also kept in a secure location.Bacterial and yeast isolates were stored according to the Standard Operating Procedure (SOP) of St. Paul’s Hospital Millennium Medical College (SPHMMC).
All data for the surveys were coded, double entered, and analyzed using the Statistical Package for the Social Sciences (SPSS) software version 23.Use descriptive statistics and logistic regression to estimate rough ratios with 95% confidence intervals for different variables.P values ​​< 0.05 were considered significant.
Urine samples were collected from each pediatric patient using sterile urine containers.Parents or guardians of study participants were given appropriate instructions on how to collect clean-captured midstream urine samples.Catheter and suprapubic urine samples were collected by trained nurses and physicians.Immediately after collection, samples were taken to SPHMMC’s microbiology laboratory for further processing.Parts of the samples were inoculated onto MacConkey agar plates (Oxoid, Basingstoke and Hampshire, England) and blood agar (Oxoid, Basingstoke and Hampshire, England) media in a safety cabinet using a 1 μL calibration loop.The remaining samples were plated on brain heart infusion agar supplemented with chloramphenicol (100 µgml-1) and gentamicin (50 µgml-1) (Oxoid, Basingstoke, and Hampshire, England).
All inoculated plates were incubated aerobically at 37°C for 18-48 hours and checked for bacterial and/or yeast growth.Colony counts of bacteria or yeast producing ≥105 cfu/mL urine were considered significant growth.Urine samples yielding three or more species were not considered for further investigation.
Pure isolates of bacterial pathogens were initially characterized by colony morphology, Gram staining.Gram-positive bacteria were further characterized using catalase, bile aescin, pyrrolidinopeptidase (PRY) and rabbit plasma.Gram-negative bacteria through routine biochemical tests such as (urease test, indole test, citrate utilization test, trisaccharide iron test, hydrogen sulfide (H2S) production test, lysine iron agar test, motility test and oxidase test test) to the species level).
Yeasts were identified using routine routine diagnostic methods such as Gram staining, embryo tube assays, carbohydrate fermentation and assimilation assays using chromogenic medium (CHROMagar Candida medium, bioM’erieux, France) according to the manufacturer’s instructions.
Antimicrobial susceptibility testing was performed by Kirby Bauer disc diffusion on Mueller Hinton agar (Oxoid, Basingstoke, England) according to Clinical Laboratory Standards Institute (CLSI) guidelines24.Bacterial suspensions of each isolate were prepared in 0.5 mL of nutrient broth and adjusted for turbidity to match the 0.5 McFarland standard to obtain approximately 1 × 106 colony-forming units (CFUs) per mL of biomass.Dip a sterile swab into the suspension and remove excess material by pressing it against the side of the tube.The swabs were then smeared in the center of a Mueller Hinton agar plate and distributed evenly over the medium.Antibiotic disks were placed on Mueller Hinton agar seeded with each isolate within 15 minutes of inoculation and incubated at 35-37 °C for 24 hours.Use a caliper to measure the diameter of the zone of inhibition.Diameter-area inhibition was interpreted as sensitive (S), intermediate (I), or resistant (R) according to Clinical and Laboratory Standards Institute (CLSI) guidelines24.Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 25922) and Pseudomonas aeruginosa (ATCC 27853) were used as quality control strains to check the efficacy of antibiotics.
For Gram-negative bacteria, we use antibiotic plates: amoxicillin/clavulanate (30 μg); ciprofloxacin (5 μg); nitrofurantoin (300 μg); ampicillin (10 μg); amikacin (30 μg); Meropenem (10 μg); Piperacillin-tazobactam (100/10 μg); Cefazolin (30 μg); Trimethoprim-sulfamethoxazole (1.25/23.75 μg).
Antibacterial discs for Gram-positive isolates were: penicillin (10 units); cefoxitin (30 μg); nitrofurantoin (300 μg); vancomycin (30 μg); trimethoprim-sulfamethoxazole (1.25/g) 23.75 μg); Ciprofloxacin (5 μg); Doxycycline (30 μg).All antimicrobial discs used in our study were products of Oxide, Basingstoke and Hampshire, England.
As shown in Table 1, this study enrolled 227 (227) pediatric patients who demonstrated or were highly suspected of having a UTI and met selection criteria.Male study participants (138; 60.8%) outnumbered female study participants (89; 39.2%), with a female to male ratio of 1.6:1.The number of study subjects was variable across age groups, with the ˂ 3-year-old age group having the most patients (119; 52.4%), followed by the 13-15-year-old (37; 16.3%) and 3-6-year-old age groups (31; 13.7%), respectively.The research objects are mainly cities, with an urban-rural ratio of 2.4:1 (Table 1).
Table 1 Socio-demographic characteristics of study subjects and frequency of culturally positive samples (N= 227)
Significant bacterial/yeast growth was observed in 65 of 227 (227) urine samples for an overall prevalence of 28.6% (65/227), of which 21.6% (49/227) were bacterial pathogens, while 7 % (16/227) were fungal pathogens.The prevalence of UTI was highest in the 13-15 year age group at 17/37 (46.0%) and in the 10-12 year age group it was lowest at 2/21 (9.5%).Table 2).Women had a higher rate of UTIs, 30/89 (33.7%), compared to 35/138 (25.4%) men.
Of the 49 bacterial isolates, 79.6% (39/49) were Enterobacteriaceae, of which Escherichia coli was the most common bacteria accounting for 42.9% (21/49) of total bacterial isolates, followed by Klebsiella pneumoniae bacteria, accounting for 34.6% (17/49) of bacterial isolates.Four (8.2%) isolates were represented by Acinetobacter, a non-fermenting Gram-negative bacillus.Gram-positive bacteria accounted for only 10.2% (5/49) of bacterial isolates, of which 3 (60.0%) were Enterococcus.Of the 16 yeast isolates, 6 (37.5%) were represented by C. albicans.Of the 26 community-acquired uropathogens, 76.9% (20/26) were Escherichia coli and Klebsiella pneumoniae.Of the 20 ward-acquired uropathogens, 15/20 were bacterial pathogens.Of the 19 ICU-acquired uropathogens, 10/19 were yeasts.Of the 65 culture-positive urine samples, 39 (60.0%) were hospital-acquired and 26 (40.0%) were community-acquired (Table 3).
Table 3 Logistic regression analysis of risk factors related to urinary tract infection in pediatric patients with SPHMMC (n = 227)
Among the 227 pediatric patients, 129 were hospitalized for less than 3 days, of which 25 (19.4%) were culture-positive, 120 were admitted to the outpatient clinic, of whom 25 (20.8%) were culture-positive, and 63 had a history of urinary tract infection. Among them, 23 (37.70%) were positive for culture, 38 were for indwelling catheter, 20 (52.6%) were positive for culture, and 71 were positive for body temperature >37.5°C, of ​​which 21 (29.6%) were positive for culture (Table 3).
Predictors of UTI were analyzed bivariately, and they had logistic regression values ​​for length of stay 3-6 months (COR 2.122; 95% CI: 3.31-3.43; P=0.002) and catheterization (COR= 3.56; 95) %CI: 1.73–7.1; P = 0.001).Multiple regression analysis was performed on bivariately significant predictors of UTI with the following logistic regression values: length of stay 3-6 months (AOR = 6.06, 95% CI: 1.99-18.4; P = 0.01) and catheterization ( AOR = 0.28; 95% CI: 0.13–0.57, P = 0.04).The length of hospital stay of 3-6 months was statistically significantly associated with UTI (P = 0.01).The association of UTI with catheterization was also statistically significant (P=0.04).However, residence, sex, age, source of admission, previous history of UTI, HIV status, body temperature, and chronic infection were not found to be significantly associated with UTI (Table 3).
Tables 4 and 5 describe the overall antimicrobial susceptibility patterns of Gram-negative and Gram-positive bacteria to the nine antibiotics evaluated.Amikacin and meropenem were the most effective drugs tested against Gram-negative bacteria, with resistance rates of 4.6% and 9.1%, respectively.Among all tested drugs, Gram-negative bacteria were the most resistant to ampicillin, cefazolin, and trimethoprim-sulfamethoxazole, with resistance rates of 100%, 92.1%, and 84.1%, respectively.E. coli, the most common recovered species, had higher resistance to ampicillin (100%), cefazolin (90.5%), and trimethoprim-sulfamethoxazole (80.0%).Klebsiella pneumoniae was the second most frequently isolated bacterium, with a resistance rate of 94.1% to cefazolin and 88.2% to trimethoprim/sulfamethoxazole Table 4.The highest overall resistance rate (100%) of Gram-positive bacteria was observed in trimethoprim/sulfamethoxazole, but all isolates of Gram-positive bacteria (100%) were susceptible to oxacillin ( table 5).
Urinary tract infections (UTIs) remain one of the most common causes of morbidity in pediatric practice.Early diagnosis of UTI in children is important because it can be an indicator of kidney abnormalities such as scarring, hypertension, and end-stage renal disease.In our study, the prevalence of urinary tract infections was 28.6%, of which 21.6% were caused by bacterial pathogens and 7% by fungal pathogens.In our study, the extent of urinary tract infections caused by bacteria was higher than the 15.9% prevalence reported in Ethiopia by Merga Duffa et al. Similarly, 27.5% et al 19 The incidence of UTIs due to yeast in Ethiopians, especially children, is unknown for our reference.This is because fungal diseases are generally considered less important than bacterial and viral diseases in Ethiopia.Therefore, the incidence of yeast-induced urinary tract infection in pediatric patients reported in this study was 7%, the first in the country.The prevalence of UTIs caused by yeast reported in our study is consistent with the prevalence of 5.2% reported in a study in children by Seifi et al.25 However, Zarei reported a prevalence of 16.5% and 19.0% – Mahmoudabad et al 26 and Alkilani et al 27 in Iran and Egypt, respectively.The higher prevalence in these two studies is not surprising since the included study subjects were ICU patients with no age preference.Differences in the prevalence of UTIs among studies may stem from differences in study design, sociodemographic characteristics of study subjects, and comorbidities.
In the current study, 60% of UTIs were hospital-acquired (intensive care unit and ward-acquired).Similar results (78.5%) were observed by Aubron et al. 28, although the prevalence of UTIs in developing countries varied by study and by region, with no regional differences in bacterial and fungal pathogens causing UTIs.The most common bacteria recovered from urine cultures were Gram-negative bacilli, mainly Escherichia coli, followed by Klebsiella pneumoniae.6,29,30 Consistent with similar earlier studies,29,30 our study also showed that Escherichia coli was the most common bacteria.Common bacteria accounted for 42.9% of total bacterial isolates, followed by Klebsiella pneumoniae, which accounted for 34.6% of bacterial isolates.Escherichia coli was the most common bacterial pathogen in community- and hospital-acquired UTIs (57.1% and 42.9%, respectively).Multiple studies have shown that Candida is the cause of at least 10-15% of hospital-acquired urinary tract infections in hospital settings, and candida is especially common in intensive care units.31-33 In our study, Candida accounted for 7% of UTIs, 94% of which were nosocomial-acquired, of which 62.5% were observed in ICU patients.Candida albicans was the main cause of candidiasis, and 81.1% of Candida were isolated from ward-acquired urine culture-positive and ICU-acquired positive urine culture samples.Our results are not surprising since Candida is an opportunistic pathogen that can cause illness in immunocompromised patients such as ICU patients.
In this study, women were more susceptible than men to urinary tract infections, and patients in the 12-15 age group were more susceptible.However, the difference between the two conditions was not statistically significant.The lack of association between UTI and gender and age can be described by the primary age group in which patients were recruited.Given the known epidemiological patterns of UTIs, the incidence of males and females generally appears to be equal in infancy, with male predominance in the neonatal period and female predominance in early childhood and during toilet training.Among other statistically analyzed risk factors, hospital stay of 3-30 days was statistically associated with UTI (P=0.01).A correlation between length of hospital stay and UTI was observed in other studies.34,35 UTI in our study was also significantly associated with catheterization (P=0.04).According to Gokula et al. 35 and Saint et al. 36, catheterization increased the threat of UTIs by 3 to 10%, depending on the length of catheterization.Sterility prevention issues during catheter insertion, infrequent catheter replacement, and poor catheter care may account for the increase in catheter-related urinary tract infections.
During the study period, more pediatric patients under three years of age were admitted to hospital with symptoms of urinary tract infection than other age groups.This may be because this age is the age for potty training, which is consistent with other studies.37-39
In this study, Gram-negative bacteria were the most resistant to ampicillin and trimethoprim-sulfamethoxazole, with resistance rates of 100% and 84.1%, respectively.The most frequently recovered Escherichia coli and Klebsiella pneumoniae were more resistant to ampicillin (100%) and trimethoprim-sulfamethoxazole (81.0%).Likewise, the highest overall resistance rate (100%) in Gram-positive bacteria was observed in trimethoprim/sulfamethoxazole.Ampicillin and trimethoprim-sulfamethoxazole have been widely used as first-line empiric treatment of urinary tract infections in all health facilities in Ethiopia, as recommended by the Ministry of Health’s Standard Treatment Guidelines (STG).40-42 Resistance rates of gram-negative and gram-positive bacteria to ampicillin and trimethoprim-sulfamethoxazole in this study.Continued use of drugs in the community increases the likelihood of selection and maintenance of resistant strains in that setting.43-45 On the other hand, our study showed that amikacin and meropenem were the most effective drugs against Gram-negative bacteria and oxacillin was the most effective drug against Gram-positive bacteria.The data in this article is taken from an unpublished paper by Nuhamen Zena, which has been uploaded to the Addis Ababa University Institutional Repository.46
Due to resource constraints, we were unable to perform antifungal susceptibility testing on the fungal pathogens identified in this study.
The overall prevalence of UTIs was 28.6%, of which 75.4% (49/65) were bacterial-related UTIs and 24.6% (19/65) were yeast-caused UTIs.Enterobacteriaceae are the leading cause of urinary tract infections.Both C. albicans and non-albicans C. albicans have been associated with yeast-induced UTIs, especially in ICU patients.Length of hospital stay and catheterization of 3 to 6 months were significantly associated with UTI.Both gram-negative and gram-positive bacteria are highly resistant to ampicillin and trimethoprim-sulfamethoxazole recommended by the Ministry of Health for empiric treatment of UTIs.Further work should be done on UTIs in children, and ampicillin and trimethoprim-sulfamethoxazole should be reconsidered as the drugs of choice for empiric treatment of UTIs.
The study was conducted in accordance with the Declaration of Helsinki.All ethical considerations and obligations were properly addressed and the research was conducted with ethical clearance and SPHMMC permission from the Internal Review Board of the Department of Medical Laboratory Sciences, Faculty of Health Sciences, Addis Ababa University.Since our study involved children (under 16 years old), they were unable to give genuine written consent.Therefore, the consent form is to be filled out by the parent/guardian.In short, the purpose of the work and its benefits are clearly described to each parent/guardian.Parents/guardians are advised that each child’s personal information will be kept confidential.The parent/guardian is informed that his or her child is under no obligation to participate in the study if he/she does not consent to participate in the study.Once they have agreed to participate in the study and are not interested in continuing, they are free to withdraw from the study at any time during the study.
We would like to thank the attending pediatrician at the study site for rigorous review of the patients from a clinical presentation perspective.We are also very grateful to the patients who participated in the study.We would also like to thank Nuhamen Zena for allowing us to extract important data from her unpublished research, which has been uploaded to the Addis Ababa University repository.
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Post time: Apr-14-2022