Biofilm Forming Capability, Multidrug Resistance and Detection of Associated Genes in Uropathogenic Escherichia coli isolated from Catheterized Patients

ZANCO Journal of Pure and Applied Sciences The official scientific journal of Salahaddin University-Erbil https://zancojournals.su.edu.krd/index.php/JPAS ISSN (print ):2218-0230, ISSN (online): 2412-3986, DOI: http://dx.doi.org/10.21271/zjpas RESEARCH PAPER Biofilm Forming Capability, Multidrug Resistance and Detection of Associated Genes in Uropathogenic Escherichia coli isolated from Catheterized Patients Iman A. Muhammad1 and Daristan J. Ghareb2 1&2 Department of Biology, College of Science, Salahaddin University-Erbil, Kurdistan Region, Iraq A B S T R A C T: Urinary tract infection (UTI) is one of the most widely recognized bacterial infections worldwide. Uropathogenic Escherichia coli (UPEC) is one of the main causative agents for nosocomial UTI. The microbial biofilms cause a public health problem to persons who require indwelling medical devices such as a urinary catheter. The pathogens in biofilms are protected from unfavorable conditions, antibiotic therapies and the host's immune system. The aim of the present study was to detect biofilm producer isolates, find out the correlation between biofilm production and associated virulence genes, and determination of antimicrobial susceptibility pattern of E. coli isolates isolated from the catheterized patient. In this study, we examined 43 isolates of E. coli collected from catheterized patients at three hospitals inside Erbil city during 25 of March to 23 of July 2018. All isolates were 100% biofilm producers using 96-well flat bottomed microtiter plate method. Also, molecular detection of uspA gene was performed as a species-specific PCR based marker for molecular identification of E. coli. fimH and papC genes were detected in 100% and 79% of the isolates respectively, as two adhesion genes responsible for biofilm formation. Minimal inhibitory concentrations (MIC50 and MIC90) were defined as the lowest concentration of antimicrobials which show 50% and 90% inhibition of planktonic form and (BIC50, BIC90) were biofilm inhibitory concentration. All isolates were 100% resistant for Ceftriaxone, Ciprofloxacin, Levofloxacin and Cotrimoxazole. While the most effective antimicrobial agent used in the current study was Nitrofurantoin. Furthermore, most of isolates in the biofilm form were more resistant to antimicrobial agents than those of the planktonic form. Additionally, there was a positive relationship between the adhesion genes and the capacity to produce biofilm in UPEC. KEY WORDS: UPEC, catheter , biofilm, adhesion gene, MIC, BIC. DOI: http://dx.doi.org/10.21271/ZJPAS.31.4.2 ZJPAS (2019) , 31(4);9-22 . 1.INTRODUCTION : Urinary tract infection is an important infection in humans, account for over 150 million cases worldwide (Terlizzi et al., 2017). Besides that UTI is the most distributed infections also reported as 36% of all health-care-associated infections (HAI) and 80% of them were expected to be catheter-associated (Parker et al., 2017) * Corresponding Author: Iman A. Muhammad E-mail: Or Article History: Received: 02/04/2019 A12/05/2019 catheter-associated urinary tract infection Accepted: Published: 10/09 (CAUTI) is/2019 defined as a UTI in patients that they have an indwelling urinary catheter at the time or within 48 hours before infection (CDC, 2018). In addition to initial urinary infection, CAUTIs can cause some other infections such as asymptomatic bacteremia, at the point when an incident of CAUTI winds up symptomatic, the subsequent sequelae can go from slight (fever, urethritis, and cystitis) to serious (calculus formation, renal damaging, acute pyelonephritis). These complications result in longer hospitalization and may increase morbidity and mortality (Niël‐ Weise and van den Broek, 2005, Flores-Mireles et al., 2015). Muhammad.I. and Ghareb.D/ZJPAS: 2019, 31 (4): 9-22 10 Escherichia coli is the most common etiologic agent of (UTIs) particularly in patients with long-term indwelling catheters. Escherichia coli is being reported to be the causative agent for more than 80% of all the UTIs and it cause both symptomatic and asymptomatic bacteriuria (Niveditha et al., 2012, Sabir et al., 2014). Pathogenicity of UPEC results from the existence and expression of many virulence factors which promote bacterial adherence and infectious expansion. Virulence genes are situated on the bacterial chromosome, plasmids or even bacteriophages and could be scattered horizontally or vertically between bacteria (Piatti et al., 2008). Biofilm is a population of cells enclosed in an extracellular polysaccharides matrix, which make them grow to biotic and abiotic surfaces (Ganjo, 2018). The extracellular matrix facilitates communications among the cells through biochemical signals, acyl-homoserine lactone in Gram-negative bacteria and oligopeptides in Gram-positive bacteria, in a phenomenon called as quorum sensing (Kim et al., 2012). The ability to form biofilm in E. coli needs several genes to be expressed which enhancing its initiation, attachment, and consequent maturation (Lee et al., 2016). A variety of virulence factors are involved in biofilm formation in E. coli, including hemolysin, fimbriae, lipopolysaccharides (LPS), secreted proteins, capsules, and ironacquisition systems, which allow attachment and bacterial colonization in the mucosal epithelial cells lining the urinary tract (Oliveira et al., 2011). In order to initiate production of a biofilm, the microorganism should initially be attached to the medical device to encourage stable attachment. The accumulation of proteins and polysaccharides in the urine favours adhesion then attachment of bacteria on to the catheter surface. When the pathogens are constantly linked to the indwelling device surface, they create exopolysaccharides to starting biofilm growth (Ohkawa et al., 1990). Two major virulence determinants of UPEC isolates are required in biofilm development: type 1 fimbriae (fim), coded by the fim gene cluster; and P-fimbriae coded by the pap (pyelonephritisassociated pili) gene (Oliveira et al., 2011). The resistance of biofilm cells to antimicrobial agents is a clinically significant feature. Bacteria forming biofilms are difficult to eliminate due to the antimicrobial resistance phenotype therefore early ZANCO Journal of Pure and Applied Sciences 2019 investigation of biofilm producers is crucial, to reduce the irrational antimicrobial burden proceeding antimicrobial resistance in the patient; then, it would be an auxiliary in controlling device-associated infections in medical clinics (Maharjan et al., 2018). The purpose of this study was to detect biofilm producer isolates, find out the correlation between biofilm production and associated virulence genes, and determination of antimicrobial susceptibility pattern of E. coli isolates isolated from the catheterized patient. 2. MATERIALS AND METHODS 2.1. Samples collection: A total of 211 urine samples were collected from catheterized patients in the following departments (urology, (...truncated)