Carbapenem resistant strains of pseudomonas species at Minia university hospitals | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Minia Journal of Medical Research | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Volume 33, Issue 4, October 2022, Page 160-165 PDF (417.41 K) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Document Type: Original Article | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
DOI: 10.21608/mjmr.2022.270692 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Authors | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Ahmed A. Saedii; Ayat M. Ahmed; Omima M. Mohamed | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Department of Clinical Pathology, Faculty of Medicine, Minia University | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Abstract | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Objectives: The incidence of Pseudomonas aeruginosa (P. aeruginosa), an important etiological cause of nosocomial infections, strains that produce beta-lactamases has been frequently recognized. Therefore, accurate and timely detection of isolates that produce carbapenemase is crucial. Aim of the work: The purpose of this study was to determine the frequency of the bla-IMP gene in carbapenem-resistant P. aeruginosa strains recovered from hospitals affiliated with Minia University. Patients and Methods: From June 2018 to September 2018, 40 P. aeruginosa strains isolated from various specimens were used in this work. Carbapenem resistant strains were gathered and analyzed for bla-IMP gene expression using PCR. Results: Out of 40 P. aeruginosa strains, 26 isolates were carbapenem resistant detected by (Vitek-2, bioMérieux, France) expressing the bla-IMP gene was in 46% of isolates. Conclusion: Carbapenem resistance in our hospitals have developed into a serious issue that calls for accurate diagnosis and efficient treatment. In this study, colistin was the antibiotic that worked the best against isolates that were resistant to carbapenem. To determine the primary source of resistance, it is advised to conduct genetic analyses for the most prevalent genes responsible for carbapenem resistance on a significant number of clinical specimens. However, given its expensive price, this might be restricted. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Conclusion Additional monitoring, robust preventative measures, and the implementation of infection-control procedures are required to stop creeping carbapenem resistance among pseudomonas strains. For all isolates, molecular validation by PCR of various carbapenemase manufacturers is necessary in addition to standard phenotypic methods for identifying carbapenemases production in order to identify the hidden genes. The most efficient treatment policies must also be determined for each area through a regular observation programs. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Keywords | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Carbapenem; bla-IMP; metallo-β-lactamases; P.aeruginosa | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Introduction P. aeruginosa is a Gram negative aerobic, extensively spread bacteria that can live in hospitals and on a range of surfaces [1]. It is an opportunistic pathogen that accounts for 10–20% of nosocomial infections, including cystic fibrosis, urinary tract infections, pneumonia, burn infections, and wound infections[2]. It can also cause bacteremia and sepsis in intensive care units. P. aeruginosa that is multidrug resistant (MDR) is resistant to at least three anti-pseudomonal antibiotics (quinolones, aminoglycosides, carbapenems, and penicillin/cephalosporin)[3]. Due to the organism's intrinsic resistance to numerous antibiotic classes and its ability to develop resistance to all potent antimicrobial medications, MDR P. aeruginosa is becoming a significant health concern [4]. P. aeruginosa produces metallo-lactamases (MβLs) enzymes, including IMP (imipenem active), which give rise to a variety of mechanisms of resistance against carbapenems. by creating the metallo-lactamases (MβLs) enzymes, particularly the IMP (imipenem activemetallo-lactamase) and VIM (Verona integron-encoded metallo-lactamase) [5, 6]. Ambler classes A, B, and D are mentioned in relation to carbapenemase resistance [7,8], although MβL VIM and IMP are classified as Ambler B. Ethylene-diamine-tetra-acetic acid (EDTA) and sodium mercapto-acetic acid (SMA) inhibit MβLs, whereas β-lactase inhibitors such clavulanic acid, sulbactam, and tazobactam had no effect on MLs [9]. The purpose of this study was to find out how frequently P. aeruginosa strains isolated from patients at Minia university hospitals included the bla-IMP gene.
Material and methods Isolation and identification P. aeruginosa The 40 pseudomonas species used in this study were obtained from clinical specimens sent to the Minia University hospitals' microbiology unit for culture and sensitivity testing between June 2018 and September 2018. The following procedures were applied to all isolates: - Routine culture on blood and MacConkey agar media for 24-48 hours incubation at 37°C. Then identification and AST was done using (VITEK-2, bioMérieux - France). - All Carbapenems (Imipenem and Meropenem) resistant pseudomonas isolates were preserved for evaluation of bla-IMP gene expression using PCR. - DNA extraction was done using (Promega Co., USA) kits following the manufacturer’s protocol. - PCR detection: particular primer created from (bla-IMP): Reverse primer: (5'-AAC CAG TTT TGC CTT ACC AT-3'), forward primer: (5'-CTA CCG CAG CAG AGT CTT TG-3'). (Germany's Operant Co.) [10]. The primer pair (5'-ATGGAAATGCTGAAATTCGGC-3') and (5'-CTTCTTCAGCTCGACGCGACG-3') was chosen as a reference for our research in order to amplify conserved portions of a target gene in P. aeruginosa and produce an identifiable PCR amplicon (500 bp) by gel electrophoresis.
Statistical analysis The means and standard deviations of the numerical data were used to express them. Frequencies and percentages were used to express the qualitative data. Results 40 Pseudomonas isolates from clinical specimens admitted to the microbiology unit at Minia university hospitals were used in this study, and they were dispersed based on the type of specimens as shown in the table (1). Table (1): Pseudomonas distribution by specimen type:
Table (2): Antibiotic resistance pattern of isolated Pseudomonas spp.:
Table (3): Distribution of carbapenem resistant Pseudomonas isolates according to the type of specimens
Table (4): Antibiotic resistance pattern of the carbapenem resistant isolates:
In this study, the resistance pattern of bla-IMP gene was expressed in 30% of all isolates representing about 46% of carbapenem resistant strains.
Figure (1): bla-IMP gene PCR detection on agarose gel electrophoresis. Samples in lanes (1, 2, 3, 4, and 5) tested positive for the bla-IMP gene.
Discussion MDR P. aeruginosa is regarded as a serious issue, and a variety of antibiotics were used, including piperacillin-tazobactam, third-generation cephalosporin (ceftazidime), fourth-generation cephalosporin (cefepime), carbape-nems (imipenem and meropenem), amino-glycosides (amikacin, gentamicin, and tobramycin), fluoroquinolones (ciprofloxacin, ofloxacin, and levofloxacin), monobactam (aztreonam), and colistin [11]. Particularly with imipenem, the antibiotic resistance pattern in pseudomonas slacks off quickly [12]. When treating MDR P. aeruginosa, carbapenems (such as imipenem and meropenem) are frequently utilized as a last option [13]. Antibiotic resistance to carbapenems is thought to be primarily caused by the production or acquisition of MβL genes (such as IMP and VIM) [14, 15]. The infection control practises and strategies followed in hospitals influence the MβLs that produce P.aeruginosa in Egypt [16]. According to the current study, Carbapenem resistance was observed in 65% of P. aeruginosa clinical isolates against (imipenem and meropenem). These outcomes were comparable to those reported by Rehab Mousa et al.,[17] who reported 69% of her isolates. In our study, P. aeruginosa had a 62.5% imipenem resistance rate and a 65% meropenem resistance rate. According on regional antibiotic policies, strain origin, and geographic location, carbapenem resistance rates can change. In this work, bla-IMP gene was expressed in 30% of all pseudomonas isolates representing about 46% of carbapenem resistant strains. Al-Agamy et al., discovered that 34% of his pseudomonas strains had carbapenem resis-tance, and roughly 22% of these isolates developed MBLs [18]. On the other hand, our findings contradict a research from Iran that claimed that only 8 (9.75%) of P. aeruginosa isolates that produced MBL were positive for bla-IMP [19]. And about 70 (70%) of the 100 imipenem-resistant pseudomonas isolates in a study conducted in Iran were identified to produce MBLs [20]. In order to provide suitable targeted therapy and prevent the nosocomial transmission of these resistant strains, it was crucial to discover MBL-producing isolates [21]. According to the results of the current work, several P. aeruginosa isolates showed varying degrees of resistance to aminoglycosides, including Amikacin and Gentamicin, with percentages of 60% and 55%, respectively. Similar to the current findings, earlier studies from Iran confirmed the high frequencies of P. aeruginosa resistance to aminoglycosides (gentamycin, amikacin) [22]. According to the current study, there was 60% and 55%, respectively, resistance to levo-floxacin and ciprofloxacin. These findings were in contrast to those made by Al.Fahadawi et al., in their study on pseudomonas isolates that showed good efficacy against ciprofloxacin and norfloxacin, where the percentages of sensitivity and resistance were, respectively, (85.3%, 76.5%) and (11.8%, 17.6%) [23]. In contrast, El-Badawy et al., showed that 42.4% of his isolates had at least one resistance against one quinolone antibiotic[24]. While P. aeru-ginosa isolates were demonstrated to be responsive to ciprofloxacin by Corona-Nakamura et al., [25] .The global spread of P. aeruginosa MDR strains may be to blame for this variation. One of the polymyxin class of antibiotics, colistin exhibits a broad spectrum of activity against the majority of Gram-negative bacteria. Colistin is increasingly often utilized in clinical settings, particularly for MDR P. aeruginosa. According to CLSI [26] colistin has a sensitive breakpoint of ≤ 2 mg/L and a resistant breakpoint of ≥ 4 mg/L against P. aeruginosa. Intriguingly, colistin proved to be the most effective antibiotic in our research because it was shown to be sensitive to all of the 26 isolates we tested for carbapenem resistance. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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