الفهرس 
Iron transport systemsيوجد فقط 14 صفحة متاحة للعرض العام
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المستخلص
The present study aimed to detect the relationship between virulence factors and antimicrobial resistance of K. pneumoniae clinical isolates. A total of 40 K. pneumoniae clinical isolates were obtained from Microbiology and Immunology Department, Faculty of pharmacy, Helwan University, Egypt. The sources of samples were urine (50 %), sputum (20 %), pus (15 %), blood (7.5 %) and throat swab (7.5 %). Hundred percentage of the tested K. pneumoniae isolates were capsulated, 90 % had fimbriae, 87.5 % were biofilm former and 17.5 % were haemolysin producer. The susceptibility of the recovered 40 K. pneumoniae clinical isolates to 33 different antimicrobials was determined using disk diffusion method and confirmed by automated Vitek2® system (BioMérieux®). The incidence of resistance to antimicrobial agents ranged from 5 % (tigecycline) to 100 % (ampicillin, amxicillin, pireracillin, ticarcillin, cephradine and cefoxitin). The resistance patterns of the tested isolates were heterogonous hence a total 16 patterns were detected among tested isolates. Each pattern was exhibited by only 1, 2 or 3 isolates. The MAR index analysis revealed that 33 (82.5 %) isolates had a very high MAR index value (≥ 0.51). These isolates were 42.5 % (MDR), 35 % (XDR) and 5 % (PDR). The high MICs values of different antimicrobials which were recorded for the tested isolates exhibited that, the MICs of ampicillin, amoxicillin, cephradine and chloramplenicol ranged from ≥ 1024 to 128 μg/ml. While MICs of ceftazidime, cefotaxime, cefoperazone, ceftriaxone and cefepime ranged from 1024 to 64 μg/ml. MICs values of carbapenems (imipenem and meropenem), fluoroquinolones (ciprofloxacin and levofloxacin) and aminoglycosides (amikacin and gentamicin) ranged from 512 to 32 μg/ml. All isolates were tested for ESBLs producing using DDDT. Out of the 40 tested isolates 40 % were ESBLs producer. ESBLs producing isolates showed high incidence (100 %) of virulence factors as fimbriae and biofilm as compared to ESBLs non producing isolates where incidence of fimbriae and biofilm were 83.3 % and 79.1 % respectively. The relationships between antimicrobial resistance and virulence factors production were studied. Our data revealed that, there was a significant association between antimicrobial resistance and biofilm formation among tested isolates. Biofilm producer K. pneumoniae isolates had high incidence of β-lactams resistance (100 % to 37.1 %), fluroquinolones resistance (57.1 % to 48.5 %), aminoglycosides resistance (40 % to 28.5), chloramphenicol (94.1 %), nitrofurantion (77.1 %), tetracycline (71.4 %) and trimethoprim/ sulfamethoxazole (62.8 %) compared to biofilm non producer isolates (all of them were sensitive to carbapenems and all other non β-lactam tested antimicrobials). There was a signification association between antimicrobial resistance and haemolysin production. All of haemolysin producer (100 %) were resistant to chloramphenicol, tetracycline, penicillins, cephalosporins and monobactams. Also, these isolates showed high incidence of resistance 85.7 % for gentamicin, tobramycin, nitrofurantion and tetracycline, 71.4 % for ciprofloxacin, gatifloxacin, levofloxacin, moxifloxacin, norfloxacin and amikacin and 42.8 % for carbapenems. There was a signification association between presence of fimbriae and antimicrobial resistance. Fimbriae producing isolates showed high incidence of resistance for β-lactams (100 % to 36.1 %), fluroquinolones (55.5 % to 47.2 %), aminoglycosides (38.8 % to 27.7 %), chloramphenicol (91.6 %), nitrofurantion (75 %), tetracycline (69.4 %) and trimethoprim/ sulfamethoxazole (61.1 %) compared to isolate lack fimbriae that were sensitive to carbapenems and all non β-lactams. All selected MDR, XDR and PDR isolates harbored 1-3 plasmids with different sizes. The genotypic detection of virulence and antimicrobial resistance genes indicated that, the most common virulence genes were mrkD (100 %) and uge (90 %) followed by FimH (50 %), irp2 and kfu (40 % for each) and GyrB2 (30 %). While, the most common resistance genes were IntI1 (80 %), CTX-M-1 (70 %) and qnrS and aacA4 (60 % for each) followed by aac(6’)-Ib-cr (50 %), TEM and qnrA (30 % for each), aacC1 (20 %) and IMP, OXA-48 and qepA (10 % for each). There was a positive relationship between presence of virulence genes and ability of isolates to resist antimicrobials. DNA sequencing of the isolates K1 (for OXA-48 gene), K7 (for OmpK- 35 gene), K8 (for irp2 gene) and K12 (for qnrA, qnrS and IntI1 genes) were carried out and the data were deposited in the GenBank databases with the following accession numbers MF490442 (IntI1), MF490443 (irp2), MF490444 (OmpK-35), MF490445 (OXA-48), MF490446 (qnrA) and MF490447 (qnrS). The DNA sequence analysis of our OXA-48, qnrS and IntI1 genes showed 100% identity with the sequence of global genotypes. Furthermore, data analysis of qnrA, irp2 and OmpK-35 DNA sequencing from our isolates showed variance in the sequence when compared to other published sequences of qnrA, irp2 and OmpK-35 using BLAST analysis. There were transversion type of mutation appeared due to frequent direct or indirect transmissions between the local area and foreign countries, indicating that our K. pneumoniae isolates are part of globally circulating lineages. To ensure the role of the detected plasmids in antimicrobial resistance and virulence of selected isolates, plasmid curing was conducted using ethidium bromide. The cured isolates lost virulence genes as GyrB2 and kfu and their resistance to some antimicrobials ranged from 4 (12.1 %) to 16 (48.5 %) antimicrobials. Loss of resistance markers and virulence genes indicated that, these virulence genes and the genes encoding resistance to those markers were located on plasmid.