Evaluation of the Possibility of Chromosomal and Plasmid-Dependent Antibiotic-Transferable Genes in Acinetobacter Genomic Isolates Isolated from Clinical Specimens and genotype

Acinetobacter baumannii is an opportunistic pathogen and a component of gram-negative, aerobic, and non-fermentative gram-negative bacteria, which is found in the form of cocci or coccobacilli. Because these bacteria have few nutrients to grow, they can survive long periods in adverse conditions, on dry surfaces, as well as in aquatic environments. Acinetobacter is probably known as gram-negative bacteria on the surface of the genus. These organisms are difficult to stain and are often mistaken for gram-positive. Among the gram-negative bacteria that cause nosocomial infections (especially in the intensive care unit of the ICU), Acinetobacter species have received a great deal of attention over the last three decades (1-3).Acinetobacter baumannii OmpA binds to the host epithelium and mitochondria, causing mitochondrial dysfunction and swelling that is followed by the release of cytochrome c, leading to the formation of apoptosis, all of which contribute to cell apoptosis. OmpA, the most abundant surface protein in the pathogen, is also involved in complement resistance and biofilm formation. Key stress survival strategies and the potential for important virulence factors help increase bacterial survival inside and outside the host (1, 4). Most chromosomal AmpC beta-lactamases are found in Enterobacter, Serachia, Pseudomonas, Acinetobacter and, Citrobacter species. These enzymes belong to the C ambler classification and include the genes cmy, fox, mox, dha, acc, mic, family / related bil / lat (cit and act, mir (related to the ebc family)). They are (5). It is noteworthy that at tigecycline resistance levels in Acinetobacter baumannii isolates, its efficacy may be increased during treatment with tigecycline if the drug is exposed for a short time. Recently, mutations in the trm gene, encoding methyltransferases, have been associated with decreased tigecycline sensitivity in one Acinetobacter baumannii strain (6). Most strains of Acinetobacter baumannii are resistant to ampicillin, amoxicillin-clavulanic acid, anti-staphylococcal penicillins, broad-spectrum cephalosporins (except ceftazidime and cefepime), tetracycline,and, tetracycline. MDR resistance in Acinetobacter strains has become a global and growing problem. In previous studies in Iran, the rate of multidrug resistance has been more than 60% (7).

A total of 240 clinical samples (including blood, urine, sputum, respiratory secretions, urine, wounds, skin, etc.) were collected from patients admitted to different wards of Dey Hospital in Tehran. Samples were collected from patients who had been hospitalized for at least three days and had acquired the infection in a hospital setting and were transferred to a microbiology laboratory for evaluation. For initial isolation, clinical specimens were cultured linearly in McConkey Agar and Bloodagar media containing 5% sheep blood and incubated at 37 ° C for 24-48 hours (9). After the incubation period, the cultures were examined for macroscopic characteristics (appearance of the colonies) and microscopy using a hot staining technique. Then gram-negative bacilli were analyzed using biochemical tests.In order to extract the plasmid, the extraction kit made by Sina Clone Company, Iran was used. First, a colony of each isolate cultured on Müller-Hinton agar medium (manufactured by Merck, Germany) was inoculated with 5 ml of Luria Bertani Broth culture medium (manufactured by Sigma-Aldrich, USA) and stored for 13 hours in Incubated at 39 ° C. Then, the plasmid extraction steps were performed using the Cinnagen kit protocol. It should be noted that the resulting plasmids were stored at -20 ° C until PCR.

According to the results in Table 6, the higher the number of isolates containing mox gene, the more antibiotic resistance is observed. In fact, a significant relationship was found between the presence of this gene and antibiotic resistance. For colistin, which had the lowest resistance ratio of this antibiotic, fewer isolates had this gene. (Significance level 0.563) A semi-sensitive to colistin isolate containing both ox mox. Genes. One of the semi-sensitive isolates containing cefpeme contained only dha and no semi-susceptible isolates contained cit gene (significance level 0.853). With 1, 5 and, 14 isolates containing dha, cit and mox genes (significance level 0.802) (Table 6).

Previous studies have shown that resistance to various antibiotics is on the rise. For example, studies by Wang et al. (23) and Esmoliako et al. (24) in recent years have shown that most strains were sensitive to amikacin, ampicillin, sulbactam, ceftazidime, cefpime, gentamicin, imipenem, meropenem, piperacillin, tazobactam. . In contrast to the present study, which found a 98.3% resistance to ampicillin and the highest resistance among all antibiotics studied, Karimi et al. reported a sensitivity of 91.6% to ampicillin in 2020. Consistent with the results of the present study, the results of the study of money changers and colleagues (2019) in Shahrekord have also shown a high prevalence of resistance to carbapenems, impenem 78% and meropenem 44%. In this study, Acinetobacter baumannii had the highest resistance to cefipime and ceftazidime (100%) and the lowest resistance to tobramycin and meropenem (22). In the study of Karimi et al., The resistance to meropenem was 83.3% and to ceftazidime was 93.3%. (19) Salehnia et al. In their study showed 100% resistance to cefpeme (7).According to the antibiogram results of the present study and also the results of the mentioned studies, to date, some strains of Acinetobacter baumannii have become resistant to all common antibiotics used, which greatly limits the treatment of these infections. The only effective antibiotic used to treat infections associated with this bacterium is colistin, which is also increasing its resistance. In general, the findings of the present study indicate an increase in antibiotic resistance compared to previous studies. This high prevalence of resistance is due to the unnecessary prescription of antibiotics and the lack of appropriate infection control tools. This is very important in identifying antibiotic resistance genes. In addition, the selection of appropriate antibiotics based on antibiogram plays an important role in treating and preventing the spread of drug resistance.The results show a high percentage of MOX genes that are directly related to antibiotic resistance, however, the presence of these genes and their relationship with antibiotic resistance need further investigation. The high resistance of Acinetobacter baumannii to antibiotics was very worrying because it is difficult to control and treat this bacterium. The only effective antibiotic used to treat infections associated with this bacterium was colistin, which is also increasingly resistant.