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Antimicrobial Resistance in Animal Pseudomonas aeroginosa Clinical Isolates
Ocak, F. and Turkyilmaz, S.
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Abstract
The results of antimicrobial treatment in Pseudomonas aeroginosa infections may depend not only on the antibiotic susceptibility of the etiologic agents, but also on the biofilm production and integron carrying capabilities of the bacteria. This study aimed to examine the relationship between antimicrobial resistance and biofilm production, biofilm related virulence genes, integron genes carried by P. aeroginosa isolates obtained from different animal clinical samples. A total of 67 P. aeroginosa isolates obtained from bovine mastitis, canine otitis and dermatitis cases were used as material. Bacterial identification was carried out using conventional methods. Qualitative (Congo Red Agar Test (CRA)) and quantitative (Microplate Test (MP)) methods were used to determine the phenotypic biofilm production capacity of the isolates. Polymerase chain reaction (PCR) was used to confirm the genus and species level identification of the isolates, and to identify biofilm-associated virulence genes and integron genes. The resistance patterns of the isolates against 12 antibiotics belonging to 6 antimicrobial families were examined using the disk diffusion method. Isolates resistant to at least three drug classes were defined as multi-drug resistant (MDR). The Chi-Square (χ2) test was used to compare the relationship between the MDR capacity of the isolates and biofilm formation, the prevalence of biofilm-associated virulence genes and the prevalence of integron genes. It was determined that 41.8% of the isolates as by qualitative method and 64.2% by the quantitative method were biofilm producers. The genes responsible for biofilm formation, ppyR, pslA and pelA, were detected in 94.0%, 83.6% and 65.7% of the isolates, respectively. 23.9% of the isolates carried the integron gene. Piperacillin tazobactam and ceftolozane tazobactam were found to be the most effective drugs against P. aeroginosa isolates studied. 28.4% of the isolates were MDR. As a result of this study, it was determined that MP was more effective than CRA in determining biofilm production phenotypically. While there was no significant relationship between MDR capacities with phenotypically biofilm formation, the prevalence of ppyR and pslA virulence genes, the relationship between the prevalence of pelA virulence gene and the presence of integron genes, was significant.
Key words: Pseudomonas aeruginosa; Multiple Antibiotic Resistance; Biofilm; Integron.
Introduction
Pseudomonas aeruginosa is a Gram-negative, opportunistic pathogen that is commonly found in soil, water, sewage and mammalian intestines, often causing infections in immuno-compromised individuals (1, 2). This opportunistic pathogen is gaining increasing importance as a causative agent of many diseases in animals, including otitis media and pyoderma in dogs and mastitis in cows (3-6). While P. aeruginosa is naturally resistant to many antimicrobial agents, it also has the capacity to rapidly develop resistance to many drugs during treatment. Therefore, the treatment of infections caused by P. aeruginosa is difficult due to both naturally occurring (intrinsic) and acquired (acquired) resistance in these bacteria (1, 2).
To survive, microorganisms have developed, various strategies such as slime production and development of biofilms to colonize and cause infections. A bacterial biofilm is a bacterial community embedded in a self-produced matrix (i.e. slime) (7). It is known that P. aeruginosa has the ability to form biofilms and that there are three important genes (pslA, pela, ppyR) that encode exopolysaccharides that play a role in biofilm formation (8, 9). The pslA and pelA genes play an important role in the formation of the carbohydrate-rich structure of the biofilm matrix. Mutations in these two genes cause a deficiency in the bacteria’s ability to form biofilms. However, following the inactivation of the ppyR gene (the putative transmembrane protein), the psl operon is suppressed (8, 9).
Genetic elements such as plasmids, transposons and integrons play an important role in increasing the antibiotic resistance. Integrons cannot move; they contain integrase (int) genes that allow the insertion of antimicrobial resistance gene cassettes between highly conserved nucleotide sequences. Integrons can capture exogenous gene cassettes. Therefore, they play an important role in the horizontalspread of antibiotic resistance genes among bacteria. Based on the differences in the amino acid sequences of the encoded integrases, integrons have been divided into five classes. The class 1 integron with the widest variety of gene cassettes is the most clinically important and has a wide distribution among resistant strains (10).
P. aeruginosa can infect both animals and humans (1, 2). Therefore, it attracts the attention of both medical doctors and veterinarians. Although virulence genes (11) and integron genes of P. aeruginosa have been well researched in human medicine in Turkey (12), there are no studies in veterinary medicine on this subject. However, there are a limited number of studies investigating the antibiotic resistance status of P. aeroginosa isolates obtained from animal clinical samples (13, 14).
The success of antibacterial treatment in P. aeroginosa isolates may be related to the biofilm formation and integron bearing of the agent. In this study, it was aimed to examine
the relationship between multiple antimicrobial resistance capacity and biofilm production, biofilm related virulence genes (ppyR, pslA and pelA) and integron genes (int1, int2, int3) carrying the status of P. aeroginosa isolates obtained from different animal clinical samples (canine otitis/pyoderma, bovine mastitis milk). It is suggested that as a result of this study, susceptibility patterns to antibiotics commonly used in clinics for the treatment of P. aeruginosa infections in our region will be determined. Thus, the findings are prososed both as an empirical antimicrobial drug selection guide and to contribute to antimicrobial resistance surveillance programs in animal health. [...]
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The Israel Journal of Veterinary Medicine is available as an open, online journal for veterinarians worldwide.
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