Three-year evaluation of the nosocomial infections in pediatrics: bacterial and fungal profile and antimicrobial resistance pattern

Background Nosocomial infections (NIs) could lead to considerably higher mortality rates, length of the hospital stays and costs, and represent a serious public health concern worldwide. Besides, the unreasonable use of antibiotics could lead to get resistant to different antibiotics and create limited therapeutic options, increased risks of treatment failure and poor patient management. The current study aimed to evaluate the prevalence and antimicrobial susceptibility of NIs in an Iranian referral pediatrics hospital during 3 years. Methods During the 3-year period, all electronic medical records of nosocomial infection episodes in hospitalized patients were retrospectively reviewed. The bacterial and fungal profile and antimicrobial susceptibility profiles of isolates recovered from different samples of patients with NIs were determined. Results In this study, a total of 718 patients with NIs was found, among which 61.3% were male (N = 440). The median age of the patients was 2.5 years (IQR: 1 month to 3 years). Klebsiella pneumonia and Candida spp. isolates were the most prevalent microorganisms (N = 125, 17.4%, N = 121, 16.9%, respectively), followed by Pseudomonas aeruginosa (N = 72, 10%) and Coagulase-negative Staphylococci (CoNS) (N = 69, 9.6%). Pseudomonas aeroginusa strains showed high sensitivity to the studied antibiotics. Acinetobacter baumannii strains displayed more than 90% resistance to the almost all antibiotics. All of the tested isolates of S. maltophilia were susceptible to Trimethoprim−sulfamethoxazole (100%) and showed high susceptibility rate to ciprofloxacin (96.4%). Vancomycin resistance was not reported in S. aureus isolates, while 64% of Enterococcus spp. was resistant to vancomycin. The rates of methicillin resistance for S. aureus and CoNS isolates were 45.5% and 85.7%, respectively. Conclusions High frequency of antimicrobial resistance to the commonly tested antibiotics is a concerning alarm. Therefore, effective infection control programs and rational antibiotic use policies should be established promptly.


Introduction
A nosocomial infection (NI) (also known as hospitalacquired infection) is a localized or a systemic infection occurring with an adverse reaction to infectious agents that develops in 48 h or more after admission [1]. NIs could lead to considerably higher mortality rates, length of the hospital stay and costs, and represent a serious public health concern worldwide [2,3] [4,5]. Nowadays, antibiotics remain the leading therapy for treating bacterial infections. However, by the unreasonable use of antibiotics, certain strains of multidrug-resistant (MDR) bacteria have emerged by selection pressure; consequently, bacteria that have been once sensitive, reemerged as resistant to different antibiotics and create limited therapeutic options, increased risks of treatment failure and poor patient management [6]. Knowledge of proper antimicrobial prescription policy of a particular setting in addition to the investigation of causative agents and their antimicrobial susceptibility profile, is essential to improve the management and reduction of the rate of NIs [7]. The aim of the current study was the evaluation of the frequency and antimicrobial susceptibility of NIs in an Iranian children medical center during three years.

Materials and methods
This cross-sectional study was carried out in the referral hospital of Children's Medical Center, Tehran, Iran between March 2017 and February 2020. Ethical approval (IR.TUMS.CHMC.REC.1399.037) was obtained from the Ethical Committee of Tehran University of Medical Sciences, Tehran, Iran.
All patients who admitted to the medical wards of Children's Medical Center, Tehran, Iran for more than 48 h and had the evidence of NIs with positive blood, wounds and sterile fluids culture of gram-positive/gram-negative bacteria and fungi were included in this study. Duplicate isolates from one patient were excluded from the study. In vitro phenotypic characterization of bacteria or fungi was carried out using standard culture and biochemical tests as described previously [8]. The disk diffusion method or minimal inhibitory concentration (MIC) was used to test each isolate for in vitro antimicrobial susceptibility based on the Clinical and Laboratory Standards Institute criteria [9].
The following antibiotics disks from MAST Categories Ltd., Merseyside, UK, were used: imipenem (10 µg), ampicillin (10 µg), cefotaxime ( Statistical analysis of the results was performed using SPSS 13.0 (SPSS Inc. Chicago, IL, USA). The results were presented as mean, frequency and standard deviation for quantitative and percentage and frequency for qualitative data.

Results
In the current study, a total of 718 patients included, among which 61.3% were male (N = 440). The median age of the patients was 2.5 years (IQR: 1 month-3 years). Among the patients, 27.2% had underlying heart disease (N = 195) and 16.3% had seizures (N = 117). Intrinsic and acquired immunodeficiency was also reported in a number of patients (N = 35, 4.9%, N = 59, 8.2%, respectively). Three hundred and eighty-four patients (53.5%) utilized catheters, and 101 of them (14.1%) had endotracheal tube during their hospitalization.
The frequency of isolated microorganisms among the studied patients based on the sources of their isolation was mentioned in Table 1. Klebsiella pneumonia and Candida spp. were the most prevalent isolates (N = 125, 17.4%, N = 121, 16.9%, respectively), followed by P. aeruginosa (N = 72, 10%) and CoNS (N = 69, 9.6%). Also, most of the samples were isolated from blood (N = 495, 69%), followed by sterile fluids (N = 165, 23%) and finally wounds (N = 58, 8%). Klebsiella pneumonia was the most frequent organism isolated from blood and wounds, and Candida spp. was the most frequent organism isolated from sterile fluids.

Discussion
In this study, we evaluated the microorganisms isolated from NIs over three consecutive years which generally had a slow decreasing trend.
The present study showed K. pneumoniae (N = 125, 17.4%), Candida spp. (N = 121, 16.9%), and P. aeruginosa (N = 72, 10%) as the most frequent microorganisms which cause NIs among the studied children. Of course other frequent NI-causing bacteria were reported in our study including CoNS (9.6%), A. baumannii (7.1%), Psuedomonas spp. (6.7%), and Enterococcus spp. (6.5%). 61% of isolated organisms were gram-negative bacteria, which was about three times more than the number of grampositive bacteria isolated in our study (22.1%).    Likewise, high rate of gram-negative bacteria was reported in Feleke et al. (53.2%) study [6]. Also unlike the study of Feleke et al. and the one reported from Jimma which mentioned S. aureus and E. coli as their the most common isolates [6,10], in the present study, K. pneumoniae was the most common bacteria isolated. Similarly, in the study accomplished by Mahmoudi et al., [14] K. pneumoniae (n = 263, 27.5%) was reported as the most frequent bacteria. In a study by Bouza et al. [11], E. coli (35.3%) was the most commonly isolated microorganism, and Klebsiella spp. were reported as 9.8% of the pathogens. Gupta et al. [12] reported that S. aureus and CoNS as the most common isolated gram-positive bacteria which is in line with our results. Nouri et al. reported the high prevalence of gram-negative bacteria (77.9%) in NIs and low prevalence of gram-positive bacteria (22.1%), exactly as ours, and the most common bacterium causing NIs among the latter was S. aureus [13].

Stenotrophomonas maltophilia
67% of isolated strains was from ICUs (mostly NICU and PICU) (N = 482), which was compatible with the results of our previous study [14]. Also, Alvares et al. reported nosocomial pneumonia as the third most common NI in their pediatric intensive care unit [2]. Candida spp. strains were isolated frequently from PICU (25%) and emergency ICU (24.4%). Surgical and ICU patients are at higher risk of rising nosocomial fungal infections [15]. In critically ill patients, the disseminated candida infections are the principal causes of morbidity and mortality both in immunocompetent and immunocompromised patients [16].
A. baumannii strains were considerably resistant to almost all tested antibiotics except for colistin (100% sensitivity), which is similar to previous studies [17,18]. Sohail et al. [19] also showed that only 0.1% of the isolated strains were resistant to colistin. The results of study reported by Vahdani et al. [20] showed antibioticresistant A. Baumannii infections with high resistant rate to ceftazidime (96%), followed by ceftizoxime (95%), ceftriaxone (93%), ciprofloxacin (85%), and trimethoprim/ sulfamethoxazole (85%). Along with the significance of MDR A. baumannii in NIs, the increasing reports of outbreaks caused by carbapenem-resistant A. baumannii in recent years have become another frightening reality [21].
In the present study, K. pneumonia strains were highly resistant to cefotaxime (95.6%), while showed 100% susceptibility to colistin, vancomycin, ampicillin, ceftazidime and clindamycin. Sensitivity to gentamycin reported as low as 37.5% among K. pneumonia strains in our study. Compared with the results of the study by Ares et al. [22], the resistance rates of isolates in the current study against studied antibiotics, especially carbapenems, were considerably high. This difference in the resistance patterns of K. pneumoniae could be due to the different prevalent clones in Iran and other countries in addition to differences in antibiotic treatment regimens in different areas [23].
All E. coli isolates tested in this study were sensitive to nitrofurantoin and colistin, while showing significant resistance to the other antibiotics compared to our previous study [17]. However, the resistance of this microorganism to imipenem (77.8% in comparison with 8%) has increased significantly compared to the mentioned study. High resistance to ampicillin has been reported in other studies, as well [24][25][26].
There are only a limited number of studies describing the S. maltophilia infection in children [31]. Treatment of nosocomial S. maltophilia infections is complicated due to high rates of antibiotic resistance [32]. We reported 100% resistant S. maltophilia isolate to gentamycin, imipenem, and penicillin (n = 1/1). However, treatment of S. maltophilia infection is difficult due to antimicrobial resistance to a variety of agents; trimethoprimsulfamethoxazol can continue to be the first choice for