Mикробиологични и генетични проучвания върху вирулентността, резистентността и молекулярната епидемиология на клинични изолати Staphylocoссus aureus
Зареждане...
Дата
2021
Автори
Циту, Вирна-Мария Ставрос
Tsitou, Virna-Maria Stavros
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ISSN на списанието
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Издател
Медицински университет - София // Medical University - Sofia
Резюме
ЦЕЛ И ЗАДАЧИ.
Цел на настоящия дисертационен труд е:
Да се проведат изследвания върху факторите на вирулентността, антибиотичната резистентност и молекулярната епидемиология на клинично значими изолати S. aureus.
Задачи:
1. Да се колекционират клинично-значими щамове S. aureus, изолирани от български пациенти с различни стафилококови инфекции.
2. Да се разработи алгоритъм за бързо откриване на S. aureus, както и MRSA директно в положителни хемокултури и / или при пунктати от абсцеси чрез откриване на mecA (кодиращ метицилинова резистентност) и специфичен ген за видова идентификация на S . aureus чрез мултиплекс PCR.
3. Да се установи чувствителността на стафилококовите изолати към подходящи за терапията антимикробни средства и механизмите, кодиращи клинично значимата резистентност към бета-лактами, макролиди и линкозамиди.
4. Да се проучи разпределението на важни фактори на вирулентност в български клинични изолати S. aureus.
5. Да препоръча адекватна терапия въз основа на анализиране чувствителността на изследваните клинични изолати през последните 5 години, особено касаеща повлияването на MRSA, доказани в инвазивни материали.
6. Да се извърши епидемиологично типизиране и да се анализира разпространението на факторите на вирулентност и основните механизми на резистентност при секвенционни типове и клонове.
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Summary.
The aim of this dissertation is to conduct investigations on virulence factors, antibiotic resistance and molecular epidemiology of clinically relevant S. aureus isolates.
Results: The new PCR algorithm developed and tested in practice for the rapid detection of S. aureus, MSSA and MRSA / CoN, directly in positive blood cultures and / or in punctures from abscesses showed excellent specificity and sensitivity. It is a remarkable and faster alternative to routine microbiological examination. A total of 558 invasive and non-invasive clinical strains S. aureus isolated during 2016-2020 from hospitals in Sofia, were studied and evaluated for the current levels of antimicrobial resistance; mechanisms of resistance to beta-lactams and macrolides, staphylococcal RAPD type and sequence type (ST) distribution. Significant levels of macrolide resistance (34.05% for erythromycin and 23.03% for clindamycin) were found in the studied S. aureus isolates from 2016 - 2020. The level of methicillin resistance (10.93%) has slightly decreased in the last five years compared to the previous 10 years. period. Resistance to gentamicin, levofloxacin, tetracycline, chloramphenicol and the trimethoprim/sulfamethoxazole combination does not exceed 10% and is insufficiently clinically relevant. New facts have been reported in this survey on the distribution of important virulence factors in S. aureus isolates of different origin and antimicrobial susceptibility. The incidence of cna and tst is 14.2% and 6.5%, respectively as of the genes of enterotoxins, they occur in the range of 4.3% to 72.6%. Combinations of three genes, sei, seg and seb, encoding superantigens are detected in more than 60% of Bulgarian staphylococcal isolates. Predominant genes in invasive S. aureus, regardless of their antimicrobial susceptibility, were discovered in various combinations: cna, tst-1, seb, seh, sec, sed, see, seg, seh and sei, while among MRSA which were isolated in 63.6% from invasive infections and mainly from hospitalized patients, dominate the following genes encoding virulence: sea, seb, sec, seg, seh and sei. It has been found, that in penicillin-resistant staphylococcal isolates the blaZ gene is detected. While in methicillin resistant only the mecA gene has been detected. We proved that resistance to macrolides and lincosamides in Bulgaria is mainly due to ermC and ermA, followed by ermB alone or in combination, and very rarely to msrA and mefA. New evidence has been discovered for association between the combination of ermA + ermC with the staphylococcal phenotype cMLS in the studied S. aureus strains. Appropriate empirical therapy is recommended for community-acquired and hospitalrelated infections, where MRSA is more common, with a focus on invasive infections. Vancomycin, tigecycline or linezolid in invasive, systemic infections. Increasing macrolide and lincosamide resistance requires this group to be administered only after proven in vitro susceptibility. Epidemiological typing by RAPD method demonstrates widespread nosocomial distribution of 24 clones and divergence of MSSA in the period 2016-2020. The dominant type A, which circulated more intensively in 2016 and 2017, showed high epidemic and invasive potential. The isolates from the newer clone I, from 2018, showed significantly higher virulence and macrolide resistance in 42.9%, encoded by ermB and ermC. 15 MRSA clones were identified by RAPD, and five clonal complexes and 11 ST types by MLST methods respectively. Among them, the dominant sequence type is ST5CC5 - a known international epidemic clone (EMRSA), corresponding to the RAPD clone a and persisting throughout the study period with high epidemic and invasive potential. Sequence type ST582CC15, corresponding to clone b, again contained strains with high virulence and epidemicity from the three hospitals. Clone c that corresponded to ST217CC22, showing significant heterogeneity, was found in only 1 hospital and in outpatients. Clone d, resp. ST8-CC8 containing only invasive HA-MRSA is also known as international EMRSA.
Conclusion: This research of virulence, level nad mechanisms of antimicrobial resistance, serotypes distribution and molecular epidemiology of clinically relevant staphylococcal isolates gives us the basis for future monitoring of evolution and genetic relation of Bulgarian S. aureus strains in comparison with the globally spread resistant clones.
Описание
Ключови думи
микробиология; микробиологична генетика; молекулярна епидемиология; грам-положителни бактерии; вирулентност; Стафилококус ауреус - генетика // , microbiology; molecular epidemiology; gram-positive bacteria; virulence; Staphylococcus aureus - genetics