Investigation of Extended Spectrum Beta-Lactamase Profiles of Klebsiella pneumoniae Isolates by Using Genotypic and Phenotypic Methods

Yıl 2024, Cilt: 4 Sayı: 3, 76 - 82, 30.09.2024

Şükrü Şen , Ufuk Hasdemir

Öz

Objectives: The aim of this study is to detect the production of extended-spectrum beta-lactamase (ESBL) in Klebsiella pneumoniae isolates isolated in our hospital using phenotypic and genotypic methods and to evaluate the performance of the newly developed colorimetric-ESBL test in our department.
Materials and Methods: Between 2018 and 2023, routine automated susceptibility testing (VITEK 2 Compact, BioMérieux, France) in our hospital detected MICs of ceftazidime and ceftriaxone that were 100 above the ESBL screening limit (>1mg/L); 20 K. pneumoniae isolates that were below (≤1mg/L) were included in the study. The ESBL positivity and negativity of the isolates was confirmed by a combination disk test according to the EUCAST recommendations. The three most common ESBL genes, blaTEM, blaSHV and blaCTX-M, were PCR-treated with specific primers. The performance of the colorimetric-ESBL test in detecting ESBL in isolates in which at least one of the ESBL genes was detected in the PCR test was evaluated. The performance of the colorimetric-ESBL test in detecting ESBL in isolates in which at least one of the ESBL genes was detected in the PCR test was evaluated. The colorimetric-ESBL test was performed in a tube containing specific concentrations of cefotaxime/ceftriaxone and phenol red as a pH indicator (*unpublished data).
Results: All 100 isolates were confirmed as ESBL-positive by combination disk test. At least one of the ESBL genes tested by PCR was detected in 97 (97%) of the isolates. The most frequently occurring ESBL gene was blaCTX-M (95.9 %), alone or in combination with another ESBL gene. The proportion of isolates carrying all three genes together was 28.9 %. Considering the PCR test results as the gold standard, 94 of 97 ESBL-PCR positive isolates (96.9 %) gave positive results in the colorimetric- ESBL test. Of the 20 ESBL-PCR negative isolates, all (100 %) were found to be ESBL-negative in the colorimetric-ESBL test.
Conclusion: Our study showed that the most common ESBL gene in our K. pneumoniae isolates was blaCTX-M and that double or triple ESBL gene combinations were present in more than half of the isolates. The sensitivity (96.9%) and specificity (100%) of the colorimetric ESBL test in detecting ESBL was found to be high, provided that it is evaluated in large-scale studies.

Anahtar Kelimeler

antibiotic resistance, colorimetric test, extended spectrum beta-lactamase, Klebsiella pneumoniae, polymerase chain reaction

Klebsiella pneumoniae İzolatlarının Genişlemiş Spektrumlu Beta-Laktamaz Profilinin Genotipik ve Fenotipik Yöntemlerle Araştırılması

Öz

Amaç: Bu çalışmanın amacı, hastanemizde izole edilen Klebsiella pneumoniae izolatlarında genişletilmiş spektrumlu beta laktamaz (GSBL) üretimini fenotipik ve genotipik yöntemlerle tespit etmek ve GSBL saptama amacıyla bölümümüzde yeni geliştirilen kolorimetrik-GSBL testinin performansını değerlendirmektir.
Yöntem: 2018-2023 yılları arasında hastanemizde rutin otomatize duyarlılık testinde (VITEK 2 Compact, BioMérieux, Fransa) seftazidim ve seftriakson MİK’leri GSBL tarama sınırının (>1mg/L) üstünde olan 100; altında (≤1mg/L) olan 20 K. pneumoniae izolatı çalışmaya dahil edilmiştir. İzolatlarda GSBL pozitifliği ve negatifliği, EUCAST önerilerine göre kombinasyon disk testi ile konfirme edilmiştir. En sıklıkla rastlanan üç GSBL geni, blaTEM, blaSHV ve blaCTX-M, yönünden spesifik primerlerle PZR’a alınmıştır. PZR testinde GSBL genlerinden en az biri saptanan izolatlarda kolorimetrik-GSBL testinin, GSBL saptamadaki performansı değerlendirilmiştir. Kolorimetrik-GSBL testi, belirli konsantrasyonlarda sefotaksim/seftriakson ve pH indikatörü olarak fenol-kırmızısı içeren bir tüpte gerçekleştirilmiştir (araştırmacıyla görüşme sonucu).
Bulgular: 100 izolatın tamamı kombinasyon disk testi ile GSBL pozitif olarak doğrulanmıştır. İzolatların %97’sinde PZR ile araştırılan GSBL genlerinden en az biri saptanmıştır. En sıklıkla rastlanan GSBL geni, tek başına veya diğer bir GSBL geni ile birlikte blaCTX-M (%95,9) oldu. Üç geni bir arada taşıyan izolatların oranı %28,9’du. Altın standart olarak PZR test sonuçlarını dikkate aldığımızda GSBL-PZR pozitif 97 izolatın 94’ü (%96,9) kolorimetrik-GSBL testi ile pozitif sonuç vermiştir. GSBL-PZR negatif 20 izolatın ise kolorimetrik-GSBL testi ile hepsi GSBL negatif bulunmuştur.
Sonuç: Çalışmamızda, K. pneumoniae izolatlarımızda en yaygın GSBL geninin blaCTX-M olduğu ve izolatların yarısından çoğunda ikili veya üçlü GSBL gen kombinasyonlarının varlığı belirlenmiştir. Kolorimetrik-GSBL testinin, GSBL saptamadaki duyarlılığının (%96,9) ve özgüllüğünün (%100), geniş kapsamlı çalışmalarda da değerlendirilmek koşuluyla, yüksek olduğunu kaydedilmiştir.

Anahtar Kelimeler

antibiyotik direnci, genişlemiş spektrumlu beta-laktamaz, Klebsiella pneumoniae, kolorimetrik test, polimeraz zincir reaksiyonu

Kaynakça

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