Volume 7, Issue 1 (February 2020)                   IJML 2020, 7(1): 35-40 | Back to browse issues page

XML Print

Download citation:
BibTeX | RIS | EndNote | Medlars | ProCite | Reference Manager | RefWorks
Send citation to:

shakib P. Prevalence of Class 1 Integron in Klebsiella Pneumoniae Isolates from Hospitals of Sanandaj, Iran. IJML. 2020; 7 (1) :35-40
URL: http://ijml.ssu.ac.ir/article-1-323-en.html
Razi Herbal Medicines Research Center, Lorestan University of Medical Sciences, Khorramabad, Iran.
Full-Text [PDF 379 kb]   (140 Downloads)     |   Abstract (HTML)  (482 Views)
Full-Text:   (110 Views)

Klebsiella pneumoniae (K. pneumoniae) is one of the gram negative rods from Enterobacteriacea family. K. pneumoniae is the major cause of nosocomial infection, septicemia, tract respiratory infection, meningitis, and urinary tract infection [1-3]. Recently, family members of Entero-bacteriaceae especially Escherichia coli (E. coli) and K. pneumoniae are found to be resistant to most antibiotics used for treatment. Resistance to antimicrobial agent such as beta-lactam antibiotics, fluoroquinolones, and aminoglycosides in these bacteria is induced by various mechanisms [4-6].
The rapid emergence of multidrug resistant Klebsiella pneumoniae (MDRKP), becoming resistant to different categories of generally used antibiotics, is a major global medical challenge [7, 8]. Integrons as mobile genetic elements are located on the chromosome
or on a plasmid in bacteria [9]. Integrons
play a main role in the dissemination of antibiotic resistance genes among different family of bacteria [10]. Classes 1, 2, and 3 integrons are most commonly identified in Gram-negative bacteria [11].
Class 1 integrons are predominantly integron-type among the antibiotic resistance clinical isolates of Enterobacteriaceae family, including K. pneumoniae [12]. Class 1 integrons consist of two conserved regions, a 3 ́-conserved segment (3 ́-CS) and a 5 ́-conserved segment (5 ́-CS), which have a physical connection to Tn402-like transposons, and an internal variable region (VR) made up of gene cassettes in tandem that encode antimicrobial resistance determinants [13, 14].
In this study, we report the prevalence of class 1 integron in K. pneumoniae isolates from hospitals of Sanandaj in Kurdistan province.
Materials and Methods
Bacterial isolates and identification
Seventy K. pneumoniae isolates were taken from different specimens including blood, urine, wound and tracheal aspirates from October 2015 to July 2016 out of the hospitals of Sanandaj.
All the isolates were identified by Gram stain and biochemical tests such as lactose fermentation, methyl red, voges proskauer, indole, citrate (IMViC), urea hydrolysis, lysine decarboxylase, H2S production, and oxidase tests [15] .
Antibiotic susceptibility pattern of clinical isolates
Antibiotic susceptibility Pattern of  antibiotic agents ceftazidime (30 μg), cefotaxime (30 μg), ciprofloxacin (5 μg), amikacin (30 μg), gentamicin (10 μg), kanamycin (10 μg), imipenem (10 μg), and co-trimoxazole (1.25+23.75 μg) (Roscoe, Denmark) were performed according to Clinical and Laboratory Standards Institute’s (CLSI) 2016 guidelines by disc diffusion method [16].
Screening for class 1 integron by polymerase chain reaction (PCR)
After DNA extraction by SinaClon Kit, all the
isolates were screened out of class 1 integron gene by PCR amplification using Corbett thermal cycler and specific primers, as described in table 1. Cycling program was as follows: initial denaturation at 94˚C for 5 min followed by 35 cycles of denaturation at 94˚C for 1 min, annealing at 56˚C for 1 min, elongation at 72˚C for 1 min, and final elongation at 72˚C for 7 min. Then, PCR products and marker-100 bp were evaluated on 1.5% agarose gels [17]. K. pneumoniae (ATCC 1029) was used as positive control for detecting class 1 integron through PCR method.
Statistical analysis
Data were analyzed by Fisher tests with STATA software program.
Antibiotic susceptibility pattern of clinical isolates
Out of 70 K. pneumoniae, 47 (67.1%), 40 (57.1%), 24 (34.3%), 27 (38.6%), 21 (30%), 28 (40%), 6 (8.6%), and 40 (57.1%) were resistant to cefotaxime, ceftazidime, ciprofloxacin, gentamycin, kanamycin, amikacin, imipenem, and co-trimoxazole, respectively. Totally, 28 isolates (40%) were detected as multidrug resistant isolates.
Screening for class 1 integron gene
Thirteen (18.5%) out of 70 K. pneumoniae isolates caring class 1 integron gene (Fig. 1).
In our study, out of (40%) 28 multidrug resistant 11 were identified to be positive for the existence of class 1 integrons.
Table 1. Sequence of primers used in this study
Gene target Sequence of primers (5′ to 3′) Amplicon size (bp( Reference
160 bp [17]

K. pneumoniae is one of the major causes of bacterial and nasocomial infections [18, 19]. During the last decades, MDRKP isolates have increased dramatically [20, 21]. multidrug resistant isolates are resistant to at least three antibiotic groups. The emergence of multidrug resistant isolates has been proposed as the most important threat in the management of nosocomial infections [22]. Diverse strategies are involved in the spread of antibiotic resistance in K. pneumoniae strains. Among them, Integrons as one of the mobile genetic elements is described as the key factor in the dissemination of these multidrug resistant clinical isolates. An integron including the gene for an integrase site (int) and for an adjacent recombination site (attI), can be situated on the bacterial chromosome or plasmid. Class 1 integrons are the most predominant and have often been described in clinical isolates K. pneumoniae [23].
 In this study, out of 13 K. pneumoniae isolates positive for class 1 integron gene, 11 isolates were multidrug resistant. We found that class 1 integron positive isolates, compared to class 1 integron negative isolates, reveal resistance to more antibiotics. However, we identified that they are not significantly associated with the presence of class 1 integrons and multidrug resistant isolates. These results are consistent with those of Derakhshan's et al. in Tehran hospitals reporting 25.8% Class 1 integron in isolates [24]. In a study in Kashan, 82.9% were identified as multidrug resistant isolates and all multidrug resistant K. pneumoniae were positive for class 1 integrons. Moreover, Firoozeh et al. repoted high frequency of integrons among multidrug resistant isolates; this could be due to the fact that integrons bear the advantage of being found in hospitals isolates [25]. In another study, conducted by Li reported 61.4% multidrug-resistant strains and 51.1% Class 1 integron in isolates [26].
Data from this study suggest a high prevalence of class 1 integrons in K. pneumoniae strains isolated from Sanandaj, Kurdistan province, Iran. Also, The presence of class I integron genes among MDRKP highlights the need for continued identification and tracking of drug resistance in health centers to decrease the spread of multidrug resistant clinical isolates.
Conflict of Interest
The authors did not declare any conflic of interests.
We are grateful to the staff of the clinical laboratory at the Be'sat, and Tohid hospitals for providing us the possibility collecting K. pneumoniae isolates used in our study.
  1.  Minarini LA, Gales AC, Palazzo IC, Darini ALC. Prevalence of community-occurring extended spectrum β-lactamase-producing Enterobacteriaceae in Brazil. Curr  microbiol.  2007; 54(5): 335-41.
  2.  Tu YC, Lu MC, Chiang MK, Huang SP, Peng HL, Chang HY, et al. Genetic requirements for Klebsiella pneumoniae-induced liver abscess in an oral infection model. Infection and immunity 2009; 77(7): 2657-671.
  3.  Tsai YK, Fung CP, Lin JC, Chen JH, Chang FY, Chen TL, et al. Klebsiella pneumoniae outer membrane porins OmpK35 and OmpK36 play roles in both antimicrobial resistance and virulence. Antimicrob Agent Chemother. 2011; 55(4): 1485-493.
  4.  Stoesser N, Batty E, Eyre D, Morgan M, Wyllie D, Del Ojo Elias C, et al. Predicting antimicrobial susceptibilities for Escherichia coli and Klebsiella pneumoniae isolates using whole genomic sequence data. J Antimicrob Chemother. 2013; 68(10): 2234-244.
  5.  Kumarasamy KK, Toleman MA, Walsh TR, Bagaria J, Butt F, Balakrishnan R, et al. Emergence of a new antibiotic resistance mechanism in India, Pakistan, and the UK: a molecular, biological, and epidemiological study. Lancet Infect Dis. 2010; 10(9): 597-602.
  6.  Harris PN, Tambyah PA, Lye DC, Mo Y, Lee TH, Yilmaz M, et al. Effect of piperacillin-tazobactam vs meropenem on 30-day mortality for patients with E coli or Klebsiella pneumoniae bloodstream infection and ceftriaxone resistance: a randomized clinical trial. JAMA. 2018; 320(10): 984-94.
  7.  Hirsch EB, Tam VH. Detection and treatment options for Klebsiella pneumoniae carbapene-mases (KPCs): an emerging cause of multidrug-resistant infection. J Antimicrob Chemother. 2010; 65(6): 1119-125.
  8.  Li J, Nation RL, Turnidge JD, Milne RW, Coulthard K, Rayner CR, et al. Colistin: the re-emerging antibiotic for multidrug-resistant Gram-negative bacterial infections. Lancet Infect Dis. 2006; 6(9): 589-601.
  9.  Fluit A, Schmitz F. Class 1 integrons, gene cassettes, mobility, and epidemiology. Euro J Clinic Microbiol Infect Dis. 1999; 18(11): 761-70.
  10.  Arakawa Y, Murakami M, Suzuki K, Ito H, Wacharotayankun R, Ohsuka S, et al. A novel integron-like element carrying the metallo-beta-lactamase gene blaIMP. Antimicrob Agent Chemother. 1995; 39(7): 1612-615.
  11.  Boucher Y, Labbate M, Koenig JE, Stokes H. Integrons: mobilizable platforms that promote genetic diversity in bacteria. Trends Microbiol. 2007; 15(7): 301-309.
  12.  Chen L, Chavda KD, Fraimow HS, Mediavilla JR, Melano RG, Jacobs MR, et al. Complete nucleotide sequences of blaKPC-4-and blaKPC-5-harboring IncN and IncX plasmids from Klebsiella pneumoniae strains isolated in New Jersey. Antimicrob Agent Chemother. 2013; 57(1): 269-76.
  13.  Chowdhury PR, Ingold A, Vanegas N, Martínez E, Merlino J, Merkier AK, et al. Dissemination of multiple drug resistance genes by class 1 integrons in Klebsiella pneumoniae isolates from four countries: a comparative study. Antimicrob Agent Chemother. 2011; 55(7): 3140-149.
  14.  Brown HJ, Stokes H, Hall RM. The integrons In0, In2, and In5 are defective transposon derivatives. J Bacteriol. 1996; 178(15): 4429-437.
  15.  Mahon CR, Lehman DC, Manuselis G. Textbook of diagnostic microbiology-e-book: Elsevier Health Sciences; 2014.
  16.  Patel J, Cockerill F, Eliopoulos G. CLSI. Performance Standards for Antimicrobial Susceptibility Testing 26th ed CLSI supplement M100S. 2016; 36(1): 1-12.
  17.  Fiett J, Baraniak A, Mrówka A, Fleischer M, Drulis-Kawa Z, Naumiuk Ł, et al. Molecular epidemiology of acquired-metallo-β-lactamase-producing bacteria in Poland. Antimicrobial agents and chemotherapy. 2006; 50(3): 880-86.
  18.  Munoz Price LS, Poirel L, Bonomo RA, Schwaber MJ, Daikos GL, Cormican M, et al. Clinical epidemiology of the global expansion of Klebsiella pneumoniae carbapenemases. The Lancet infectious diseases. 2013; 13(9): 785-96.
  19.  Diancourt L, Passet V, Verhoef J, Grimont PA, Brisse S. Multilocus sequence typing of Klebsiella pneumoniae nosocomial isolates. J clinic microbiol. 2005; 43(8): 4178-182.
  20.  Wang X, Wang Y, Zhou Y, Li J, Yin W, Wang S, et al. Emergence of a novel mobile colistin resistance gene, mcr-8, in NDM-producing Klebsiella pneumoniae. Emerg Microb Infect. 2018; 7(1): 122.
  21.  Gu D, Dong N, Zheng Z, Lin D, Huang M, Wang L, et al. A fatal outbreak of ST11 carbapenem-resistant hypervirulent Klebsiella pneumoniae in a Chinese hospital: a molecular epidemiological study. Lancet Infect Dis. 2018; 18(1): 37-46.
  22.  Michalopoulos A, Virtzili S, Rafailidis P, Chalevelakis G, Damala M, Falagas M. Intravenous fosfomycin for the treatment of nosocomial infections caused by carbapenem resistant Klebsiella pneumoniae in critically ill patients: a prospective evaluation. Clin Microbiol nd Infect. 2010; 16(2): 184-86.
  23.  Collis CM, Kim MJ, Partridge SR, Stokes H, Hall RM. Characterization of the class 3 integron and the site-specific recombination system it determines. J Bacteriol. 2002; 184(11): 3017-3026.
  24.  Derakhshan S, Peerayeh SN, Fallah F, Bakhshi B, Rahbar M, Ashrafi A. Detection of class 1, 2, and 3 integrons among Klebsiella pneumoniae isolated from children in Tehran hospitals. Archives of Pediatric Infectious Diseases. 2014; 2(1): 164-68.
  25.  Firoozeh F, Mahluji Z, Khorshidi A, Zibaei M. Molecular characterization of class 1, 2 and 3 integrons in clinical multi-drug resistant Klebsiella pneumoniae isolates. Antimicrob Resist Infect Control 2019; 8(1): 59.
  26.  Li B, Hu Y, Wang Q, Yi Y, Woo PC, Jing H, et al. Structural diversity of class 1 integrons and their associated gene cassettes in Klebsiella pneumoniae isolates from a hospital in China. PloS One 2013; 8(9): 75805.
Type of Study: Research | Subject: Bactriology
Received: 2019/07/31 | Accepted: 2019/11/10 | Published: 2020/01/30

Add your comments about this article : Your username or Email:

Send email to the article author

© 2021 CC BY-NC 4.0 | International Journal of Medical Laboratory

Designed & Developed by : Yektaweb