Investigation of In Vitro Activity of Mecillinam Against Isolates of Enterobacteriaceae

Introduction: Mecillinam is an oral antibiotic with excellent clinical efficacy in the treatment of uncomplicated UTIs. It is a betalactam antibiotic and active against Enterobactericeae but has little activity against Gram positive bacteria. In this study, we aimed to investigate the in vitro activity of mecillinam against Enterobactericeae isolates that were isolated from different clinical specimens. Materials and methods: Identification of the bacterial isolates were tested by conventional methods and Phoenix (Becton Dickinson, USA) and Vitek MS (Biomeriux, France) automated systems. Antimicrobial susceptibility was performed by using the disk diffusion method and Phoenix (Becton Dickinson, USA) and Vitek2 Compakt (Biomeriux, France) autometed systems. Mecillinam susceptibility of the isolates were determined by disc diffusion method by using 10 μg mecillianm disc and interpreted according to the EUCAST criteria. Results: Total of 338 Enterobactericeae (E. coli n=162, Klebsiella spp. n=142, Enterobacter spp. n=18, Proteus spp. n=10, Citrobacter spp. n=4, Salmonella spp. n=2) isolates were tested. Most of the clinical specimens were urinary specimens (n=272, 80.5%). Mecillinam susceptibility was detected as 82.5%. Mecillinam susceptibility for urinary tract isolates was 87.1%. In this study, 57% of imipenem resistant isolates were determined susceptible for mecillinam. Conclusion: In conclusion, Mecillinam could be a good option, especially for urinary tract infections.


Introduction
Mecillinam is an antibiotic from beta-lactam group.Contrary to penicillin and cephalosporins, instead of 6-amidino, its structure contains 6-acylamino side chains.It is effective against Gram negative bacteria, particularly from Enterobactericeae family, while its effect against Gram positive bacteria is low [1].Upon examining the efficiency of mecillinam, it was observed that no enzyme participating in the synthesis of cell wall was inhibited and that spheroplasts with rapid lysis were not formed.It is considered that the efficiency of mecillinam comes from its affinity to penicillin-binding protein 2. Contrary to the majority of other beta-lactam antibiotics, it does not bind with penicillin-binding protein 1 or 3 [2,3].Also, due to its resistance to the effect of beta-lactamase hydrolysis, it has more effect on Enterobactericeae [4].When taken orally, 21% of the mecillinam dose is disposed of urinary (three inactive metabolites and one active metabolite), while an important amount is released through the biliary tract.
Mecillinam, especially in North European countries, is a preferred choice in the treatment of non-complicated infections of lower urinary tract because of its high-tolerability and low side effects profile [5].Mecillinam is not the preferred and frequently used antibacterial agent in our country.However, because of the rapidly increasing resistance today, we believe that in the following years it may become a good choice for treating non-complicated infections of lower urinary tract.Susceptibility of gram negative bacteria to mecillinam has not been studied in our country.In this study, our goal is not just to examine urinary system parameters, but also to examine in vitro effectiveness of mecillinam on gram negative bacteria isolated from other clinical examples, thus providing a contribution to the literature.

Material and Methods
In the study, 338 Enterobactericeae isolated from clinical samples was tested.The identification of bacteria was conducted using conventional methods (EMB Agar colony view, oxidase reaction, TSI, Indole), and urea, motion and citrate reproduction properties using Pheonix (Becton Dickinson, USA) and Vitek MS (Biomeriux, France) automatized systems.Antimicrobial susceptibility of bacterial isolates was examined with disk diffusion method using Pheoenix (Becton Dickinson, USA) and Vitek2 Compakt (Biomeriux, France) automatized system.Susceptibility to mecillinam was examined using 10 µg mecillinam disk and MHA medium with KirbyBauer disk diffusion method.The results were evaluated according to susceptibility limit values (≥15 susceptible, 15> resistant) proposed for E. coli, Klebsiella spp.and Proteus mirabilis isolated from EUCAST urinary system samples [6].

Discussion
Fosfomycin trometamol, mecillinam and nitrofurantoin are recommended the first for the treatment of noncomplicated urinary tract infections.Also, if the resistance rate to trimethoprim is not over 20%, trimethoprimsulfamethoxazole can be recommended for the first stage of the treatment as well [7].The bacteria most frequently isolated from infections of urinary tract is E. coli (80%).In the conducted studies it has been determined that E. coli is highly resistant to ampicillin, sulfamethoxazole, trimethoprim and trimethoprim-sulfamethoxazole antibiotics.
Even though mecillinam is not preferred in our country, it is the preferred agent for treating infections of urinary tract around the world, especially in North European countries.For this reason, it is advised to use mecillinam, nitrofurantoin and phosphomycin for the treatment of infections of urinary tract caused by E. coli [8].In the study by Wootton et al, the resistance to mecillinam of E. coli isolates was determined to be 6.5% [9].It was determined that the resistance to mecillinam of E. coli isolates obtained from community-onset urinary tract infections from five European countries (Austria, Portugal, Greece, United Kingdom and Sweden) was between 0-1,4%.The highest resistance rates with 1.4% were determined in Greece and Portugal [8].In our study, the resistance of E. coli isolates to mecillinam was determined to be 6%.This resistance rate is rather low, even if it is slightly higher than the resistance rates in other European countries.
In a study conducted in Canada, 2000 urinary tract isolates' susceptibility to ampicillin, trimethoprimsulfamethoxazole, mecillinam, nitrofurantoin and ciprofloxacin was examined and the resistance rates were 41.1%, 19.2%, 14.7%, 5.0% and 1.8% respectively.In the same study, it was determined that a significant portion of E. coli isolates resistant to ampicillin, trimethoprim-sulfamethoxazole or ciprofloxacin were susceptible to mecillinam [10].
In the study by Mazzuli et al. [4] susceptibility to mecillinam was examined on gram negative bacteria isolated from urinary tract infections and the susceptibility of E. coli, Klebsiella spp., Enterobacter spp., Proteus spp, and P. aeruginosa was 99.7%, 100%, 100%, 97.6% and 57.1% respectively.It was determined in the same study that the activity of mecillinam is stronger than the activity of ampillicin, trimethoprim-sulfamethoxazole and nitrofurantion, but weaker than the activity of ciprofloxacin [4].
When compared to other groups of beta-lactam antimicrobials, it was observed that mecillinam exhibited more effectiveness and stability than beta-lactamase producing isolates such as TEM, IRT and AmpC [11].There are studies that demonstrate the effectiveness of mecillinam against isolates with various resistance determinants [1,12,13].In a study conducted in India on the subject of extended spectrum beta-lactamase positive (GSBL) E. coli isolates' susceptibility to mecillinam, mecillinam resistance rate was 4.5% [12].Also, mecillinam exhibited low minimum inhibitory concentration values in GSBL positive isolates and its efficiency increased further with betalactamase inhibitory combinations [1].Since mecillinam resistance rate of NDM-1 positive E. coli isolates was determined to be as low as around 3,5%, it is believed that mecillinam can be a good choice for treatment of infections caused by this bacteria [13] .As for our study, it was observed that 57% of carbapenem-resistant isolates were susceptible to mecillinam.
Mecillinam in vitro efficiency was examined in this study on sample types other than urine samples, but on a limited scale.High mecillinam susceptibility rates were observed, except in urine isolates, especially in bacterias isolated from blood culture.Even though mecillinam has been widely used in Scandinavian countries for a long time now, resistance increase has not been observed and despite its use as only medicine for treatment of urinary tract infections, it did not cause resistance increase [14].It should be remarked that in our study over 90% susceptibility rate was determined for E. coli isolates, one of the most frequent causes for urinary tract infections.We believe that, as the number of related studies increase and mecillinam enters clinical use, the effect of this antibiotic on the isolates in our country will be understood better.

Table 1 :
Susceptibility of mecillinam according to specimens.

Table 3 :
Susceptibility of mecillinam in urine isolates resistant to amoxicillin-clavulanic acid, nitrofurantoin and

Table 4 :
Susceptibility of mecillinam in imipenem-resistant isolates