Rapid Detection of Carbapenemase Detection by VITEK-MS

Öz

In recent years, the increasing of infections caused by carbapenemase-producing Enterobacteriaceae (CPE) is an important public health problem in the worldwide. Some researchers have been reported studies showing that MALDI-TOF MS system can also be used in antibiotic susceptibility testing. The aim of this study was to evaluate the performance of VITEK-MS (bioMérieux, France) system based on ertapenem hidrolysis for the detection of carbapenemase activity. One hundered-six Klebsiella pneumoniae strains isolated from various clinical samples were tested in this study, which included 86 carbapenem resistant and 20 carbapenem susceptible isolates. The presence of KPC, NDM, IMP, VIM ve OXA-48 genes were detected by PCR. After the 4 McFarland bacterial suspensions was treated with 0.5 mg/ml of ertapenem, the suspension was incubated for 2h and 4h, and transferred to VITEK-MS RUO (bioMérieux, France) system for analysis. Eighty-six carbapenem resistant K.pneumoniae strains had blaOXA-48 (n=39), blaNDM (n= 37), blaIMP (n=8) ve blaKPC (n=2) genes. No carbapenemase gene was found in the carbapenem susceptible K.pneumoniae strains. According to results of VITEK MS, ertapenem hydrolysis was observed in all of the NDM, IMP, and KPC after 2h of incubation. Poor hydrolysis was observed in 14 OXA-48 positive strains within 2h. After 4h of incubation strong ertapenem hydrolysis was observed in 64.1 % of the OXA-48 positive strains. All non-carbapenemase producing strains were detected as negative. According to our study results, the detection of NDM, IMP and KPC using VITEK-MS seems to be a rapid, reliable and cost-effective method for routine laboratory. Since similar results were not observed in OXA-48, different protocols should be developed for the detection of these enzymes. The detection of carbapenemases by MALDI-TOF-MS systems in a short time will make a significant contribution especially in controlling the outbreaks.

Anahtar Kelimeler

• carbapenemase
• ertapenem
• Klebsiella pneumoniae
• MALDI-TOF

VITEK-MS Sistemi ile Karbapenemaz Üretiminin Hızlı Tespiti*

Öz

Son yıllarda karbapenemaz üreten Enterobacteriaceae kaynaklı enfeksiyonların dünya çapında artması önemli halk sağlığı sorunudur. Araştırıcılar, mikroorganizmanın kısa sürede tanımlanmasına olanak sağlayan MALDI-TOF sisteminin antibiyotik duyarlılık testlerinde de kullanılabileceğini gösteren çalışmalar bildirilmektedir. Çalışmamızda karbapenemaz üretiminin hızlı saptanmasına yönelik olarak ertapenemin karbapenemazlar tarafından hidrolizinin VITEK-MS (bioMérieux, Fransa) sistemi tarafından tespit edilebilirliği değerlendirilmiştir. Çalışmamıza çeşitli klinik örneklerden izole edilen karbapenem dirençli (n=86) ve duyarlı (n=20) toplam 106 Klebsiella pneumoniae kökeni dahil edilmiştir. KPC, NDM, IMP, VIM ve OXA-48 genlerinin varlığını PCR ile tespit edilmiştir. Bakteri süspansiyonları 4 McFarland bulanıklığında hazırlanmış, 0.5 µg/ml’lik ertapenem ile muamele edildikten sonra iki ve dört saat inkübe edilip analiz edilmek üzere VITEK-MS RUO (bioMérieux, Fransa) sistemine aktarılmıştır. Karbapenem dirençli 86 K.pneumoniae kökeninin blaOXA-48 (n=39), blaNDM (n= 37), blaIMP (n=8) ve blaKPC (n=2) karbapenemaz genlerini taşıdığı tespit edilmiştir. Tüm antibiyotiklere duyarlı olan kökenlerde hiçbir karbapenemaz genine rastlanmamıştır. Ertapenemin tek başına sahip olduğu spektrumlar çalışma kökenlerinin ertapenem ile muamele edilip inkübe edilmesi sonucu elde edilen spektrumlar ile karşılaştırıldığında; iki saatlik inkübasyon sonrasında NDM, IMP ve KPC geni pozitif kökenlerin tamamında ertapenem hidrolizi gözlenirken, OXA-48 geni pozitif olan 14 kökende ertapenem hidrolizinin zayıf olduğu tespit edilmiştir. Dört saatlik inkübasyon sonrası OXA-48 geni pozitif olan kökenlerin sadece % 64.1’inde ertapenem hidrolizi gözlemlenmiştir. İki ve dört saat inkübasyon sonrası karbapenemaz geni içermeyen kökenlerin hiçbirinde ertapenem hidrolizi gözlemlenmemiştir. Rutin laboratuvarda mikroorganizmaların hızlı tespitinde önemli başarı sağlayan MALDI-TOF sistemlerinin antibiyotik duyarlılık testlerinde de benzer başarı göstermesi oldukça ümit vericidir. Ancak OXA-48 pozitif kökenlerin saptanmasını kolaylaştıracak farklı protokollerin geliştirilmesi gereklidir. MALDI-TOF sistemleri ile kısa sürede, düşük maliyetle karbapenemaz üretiminin tespit edilmesi özellikle salgınların çok daha kısa sürede kontrol altına alınmasında önemli katkı sağlayacaktır.

Anahtar Kelimeler

• ertapenem
• karbapenemaz
• Klebsiella pneumoniae
• MALDI-TOF

Kaynakça

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