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DOI: http://dx.doi.org/10.20534/ESR-16-11.12-58-60
Iskhakova Khalida Ilhamovna, Tashkent Institute of Postgraduate Medical Education, Department of Microbiology MD, Professor Shadmanova Nargiza Abitovna, Head of the Department of Microbiology Tashkent Institute of Postgraduate Medical Education E-mail: shadmanova06@yahoo.com Rasulmuhamedova Munira, 1-th City Hospital of Tashkent, Head of the bacteriological laboratory
Antibiotic resistance of hospital strains of Enterobacteriaceae and phenotypic methods for detecting beta-lactamases
Abstract: Resistance of hospital infections of Enterobacteriaceae to antibiotics is important, which is not enough explored in Uzbekistan. The identification of the cultures and determination of bacterial resistance realized by conventional phenotypical methods. The high resistance of hospital strains of E. coli to cephalosporins 3-4 generations, ertapenem, and aztreonam. Highest sensitivity of E.coli indicators identified in relation to fosfomycin, netilmicin and tigecycline. Enterobacteriaceae resistant to ertapenem in phenotypic confirmatory method ICM found that 21.9% of strains were negative in tests on carbapenemases; from positive to test most frequently (in 68.0%) was the simultaneous production of MBL and KPC. It rarely detected MBL (28%), and only 4% KPC products.
Keywords: Hospital infections, antibiotic resistance, Enterobacteriaceae, beta-lactamases.
To date, fi-lactams are the most widely used antibiotics. Among them, the most popular at the moment are the cephalosporins 2-3rd generations and carbapenems (imipenem, me-ropenem and ertapenem), forming the basis of modern chemotherapy [4; 13; 15; 16; 17]. Resistance Gram-negative bacteria to p-lactam antibiotics most often associated with the production of fi-lactamases are the greatest source of resistance to them. Among the wide variety of beta-lactamases are of particular importance: 1.ESBL-extended-spectrum fi-lactamase capable of destroying all p-lactam antibiotics except carbapenems and aztreonam. 2.Car-bapenemases — enzymes, heterogeneous structure, belonging to different molecular classes, different genetic groups within groups — various genovariants. Among them, an important role is played by KPC- serine carbapenemases Class A; MBL — metallo-p-lactamases, MBL within the group the most frequent genetic groups are VIM, IMP and NDM -1. The genes responsible for the production of beta-lactamases usually located on mobile genetic elements -integrons, plasmids, and others [3; 4; 11; 13]. Detection of resistance genes in the PCR is the most reliable and accurate method, but in the daily work of microbiologist difficult doable. In the literature widely report of phenotypic methods available in the routine laboratory for detecting resistance mechanisms of bacteria to antibiotics and, in particular — the detection of various embodiments p-lactamases [5; 6; 9; 12; 14]. Phenotypic methods are important not only for the rational and competent treatment of patients with infections, they are necessary for early detection of carriers of multiresistant strains, especially in hospitalized persons (individual risk), which may contribute to the early detection of the circulation of resistant strains and timely adoption of measures to control the infection.
Purpose: Identify resistance of hospital strains of Enterobacteriaceae to p-lactam and other antibiotics, determine the frequency ofvarious products beta lactamase by phenotypic methods.
Material and methods. All strains of Enterobacteriaceae isolated from various biomaterials (pus, urine, sputum, etc.). Collection of material were performed by specialists of bacteriological laboratory of General Hospital № 1 of Tashkent and its primary selection made by conventional methods for identification of pathogens using manual Berg's [1]. Them identification and controlled by laboratory TIPME. All of strains resistance detected to 3-4th generation cephalosporin's by conventional disc-diffusion method (DDM), when assessing the limit values of the inhibition zones (LVIZ) using the recommendations of the European Committee for the sensitivity definition to antibiotics EUCAST [8] and created on the basis of this document Clinical guidelines of "Determination of the sensitivity of microorganisms to antimicrobial agents", the RF [3]. Testing was performed on Mueller-Hinton medium with commercial discs of antibiotics. For confirm the production of different variants of carbapenemases used method of combined disk — MCD [2, 5]. The method is as follows: on the disk surface of the medium overlaps with an inoculum culture studied with conventional antibiotic concentration and at a distance from it a combined drive with the same antibiotic and inhibitor. To determine the ESBL used commercial discs with cephalosporin's of the 3-th generation (ceftazidime, ceftriaxone and cefotaxime) + clavulanic acid; to determine the MBL — carbapenems antibiotic (meropenem, imipenem, ertapenem) + ETDA (Ethylenediaminetetraacetic acid); to determine the KPC — the same antibiotics carbapenems + aminophenylboronic acid; to determine the AMpC + loss of porins — the same antibiotics carbapenems + cloxacillin. Differences breakpoints around of antibiotic disc (cephalosporins or carbapenems) and disks with the same antibiotic and an inhibitor of 5 mm or more show production of the enzyme. During the observed period (2015 (0,5year) — 2016) was allocated 144 cultures of E. coli and strain 81, which consists of the combined group genera Klebsiella-Enterobacter. Single of strains of Proteus, Serratia, Citrobakter et al. did not take account of.
Antibiotic resistance of hospital strains of Enterobacteriaceae and phenotypic methods for detecting beta-lactamases
Results and discussion. Of the 144 strains of E.coli, 40 (27.8%) were positive in the preliminary screening (resistant to one of antibiotics); in Klebsiella-Enterobacter group such strains were much smaller — only 9 (11.1%) isolate from 81. Of the 49 strains of Enterobacteriaceae only 10.2% were resistant to 3-th generation of cephalosporin's, them according to EUCAST [8] and numerous other sources, presumably can be attributed to the producers of ESBL with no other enzymes that hydrolyze carbapenems. But in parallel with this, almost all Enterobacteriaceae (89.8%) were resistant to ertapenem, which raised doubts to antibiotics disks. When checking the activity of commercial discs of ertapenem with 30 mkg, microorganisms of different taxonomic groups (including the reference strains of S. aureus ATCC 25923, E.coli ATCC 25922, P.aeruginosa ATCC 27853) revealed that the breakpoints of the studied species of microorganisms (for example, S. aureus ATCC 25923-30 mm). His indicates that an almost complete absence of enterobacteria delay zones around the disc ertapenem growth is not related to the quality of the discs themselves.
Subjected to further study the most numerous group of multidrug-resistant strains of E.coli hospital, the collection was supplemented and consisted of 57 strains. The DDM determined antibiotic resistance added cefuroxime, cefepime and aztreonam; also investigated tetracyclines, chloramphenicol, fosfomycin, aminoglycosides and fluoroquinolones. The results showed a very high level of resistant to all hospital Escherichia studied cephalosporins. Share sensitive Escherichia in descending order: ceftriaxone and cefotaxime at 8.8%, 5.3% cefuroxime, cefepime 1.8% and the total lack of sensitivity to ceftazidime (moderately resistant strain of 4-7.0%). Among the most active were the carbapenems imipenem (87.7%) to meropenem sensitive strains accounted for only 54.5%, but there was a high proportion of moderately stable among the rest (1628%). Preliminary screening results on the resistance E.coli were confirmed to ertapenem — only 5 (8.8%) isolates were susceptible, resistant 91.2%. Unusual was almost complete lack of sensitivity to aztreonam E.coli only 2 of 57 (3.5%), while it is known that the antibiotic monobaktam highly active against to enterobacteria and clinics in conditions recommended as reserve medication. Among non betalaktam the greatest impact on the studied strains had fosfomycin and tigecycline — 91.2% and 86.0% respectively sensitive. Fluoro-quinolones are not highly active, there was the highest percentage sensitive to ofloxacin — 24.6%, still less to ciprofloxacin (21.1%) and levofloxacin (17.5%). Of the two studied aminoglycosides (gen-tamicin and netilmicin) was more active netilmicin — 77,2% E.coli were susceptible to it. Chloramphenicol inhibited growth of 33.3% of the strains. New breakpoints [3], according to which we carried out the study, are often different from those adopted previously [7], and the interpretation of the studied isolates breakpoints must often fall into the category of "stable." Given that our E.coli strains in a high percentage of cases have proven to be "unusual" phenotypes against carbapenems and aztreonam, we checked how the breakpoints expansion affect the strain category (sensitive, moderately resistant and resistant). As shown by these results, if you keep a record on the applicable standards before the number susceptible to merope-nem increase to 84.2%, mainly due to the transition moderately resistant to susceptible. To imipenem, ertapenem, and aztreonam categorization changes are insignificant.
Thus, almost 100% ofEnterobacteriaceae resistant to ertapenem hospital with sensitivity to imipenem and, to a lesser extent — to meropenem — these are cases that require special attention. According to [1; 8], resistance to meropenem and/or imipenem any Enterobacteriaceae (except Proteae) classifies them as "exceptional"
phenotypes, but with the caveat "except for the countries in which carbapenemases-producing Enterobacteriaceae are not uncommon." However, there is the exclusive local resistance phenotypes ofEnterobacteriaceae to ertapenem is not clear. A similar question arises with respect to aztreonam, showed an almost complete lack of activity against local strains of Escherichia. Because in our country resistant to ertapenem and aztreonam had not previously been studied, perhaps carbapenemases-producing enterobacteria are not unusual for our region. The high efficiency of ertapenem in infections caused by enterobacteria evidenced publication Kozlov R. S. et al. [10]. The great interest in this issue is due also to the fact that ertapenem is not registered in our country and in clinical practice is not used. The advantages of the product include the ability to single dose, because it is actively bound to plasma proteins, thereby increasing its half-life, and thus can receive a one-time [10]. It is also important that ertape-nem is active against ESBL-producing bacteria, which is extremely widespread in many countries of the world. Given the major role of ertapenem as a reserve drug, further work on the phenotypic identification ofproducts carbapenemases we conducted with 32 strains of Enterobacteriaceae resistant to ertapenem -24 strain of E.coli, 5 — Enterobacter cloaceae and 3 — Klebsiella pneumoniae. Each culture is resistant to ertapenem was tested in the MCD with ertapenem and combined drives to determine the MBL, cattle and AMC. As already noted, excess breakpoints diameter 5 mm or more around the disk in combination with a conventional comparison indicates the presence of a particular isolate the enzyme. The results of these studies demonstrated the following: 7 (21.9%) were negative in strains testing carbapenemases MBL, cattle or AMC, as compared with conventional disk drives combined formed around almost the same area of non-growth. 7 strains (25 of a positive testing carbapenemases) — 4 E.coli, Enterobacter cloaceae 2 and 1-Klebsiella pneumoniae, were confirmed phenotypically as producers ofMBL (28.0%). These cultures zone around the disk ertapenem ranged from 9 to 12mm, ertapenem + boronic acid 5 from 8 mm to 13 mm, while the area around the combination ertapenem + ETDA was in all cases more than 18-20 mm. 1 (4.0%) strain of Klebsiella were confirmed as producing cattle. 17 (68.0%) strains were phenotypically confirmed as producers simultaneously two carbapenemases — MBL and cattle. This primarily refers to the strains that do not respond to ertapenem (breakpoint- 0), t. E., Even at low LVIZ 13-17 mm around the combined disc, they were regarded as positive for the production of the enzyme. No isolate could not be regarded as a producer of AMC — the area around the drive with ertapenem + cloxacillin for the most part absent or did not exceed 8-10 mm under the same terms in the control disk with ertapenem.
Conclusions. 1. The high resistance of hospital strains of E. coli to cephalosporins 3-4 generations, ertapenem, and aztreonam. Highest sensitivity of E.coli indicators identified in relation to fosfo-mycin, netilmicin and tigecycline.
2. Enterobacteriaceae resistant to ertapenem in phenotypic confirmatory method ICM found that 21.9% of strains were negative in tests on carbapenemases; from positive to test most frequently (in 68.0%) met the simultaneous production of MBL and cattle producers only rarely detected MBL (28%), and only 4% was set isolated cattle products.
This report is preliminary, it is necessary to further explore other medical institutions of the city, and perhaps some region of the country. DDM must be supplemented by the definition of the minimum inhibitory concentration and, most importantly, spend genotypic study ofunusual phenotypes enterobacteria for the presence of genes encoding resistance to carbapenems.
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DOI: http://dx.doi.org/10.20534/ESR-16-11.12-60-63
Karimov Murodullo Yuldashovich, Salokhiddinov Fakhriddin Bakhriddinovich Tashkent Medical Academy Department of "Traumatology-Orthopedics, Neurosurgery № 1" E-mail: m.karimov@mail.ru, fb.doc@mail.ru,
The experience treatment by the external fixation device of our design
Abstract: The apparatus for osteosynthesis of long bone fractures has several advantages comparing to the traditional treatment methods. The rod device is easy to use and comfortable for patients. We evaluated the efficiency of the rod device in patients with fractures of the long bones.
For the period from 2011 to 2015 25 patients with fractures of the femur and tibia with multiple and associated injuries were treated using the rod device of our design. The average age of patients was 43.4 years (range 19 to 68).
The results of the study investigated in all patients from 12 months to 26 months. The average period of fixation by the device depended on the appearance of signs of consolidation and the nature of fractures. The average term for type A was 12-14 weeks, for type B and C — 14-16 months (according to AO classification). Complete fusion was observed in 22 patients.
One patient had a bilateral fracture of the shin bone, which was not observed seam leg bones, this time treated. The second patient had improper splice of shin bone as a result of early device removal. Inflammation of soft tissue around the bone rod was observed in 3 (12%) cases, which managed by subcutaneous antibiotic injections around bone rods and frequent dressings.
The designed transosseus apparatus for osteosynthesis of long bone fractures on the basis of modern locks may be the method of choice.
