Leishmaniasis şüpheli örneklerin kültür ve PCR sonuçlarının değerlendirilmesi

Year 2019, Volume: 76 Issue: 3, 313 - 320, 01.09.2019

Selma Usluca

Abstract

Amaç: Bu çalışmada leishmaniasis şüphesi ile laboratuvarımıza gönderilen kan, kemik iliği ve/veya doku biyopsisi ve yara aspiratı örneklerinin NovyMacNeal-Nicolle NNN besiyerine ekimi ve moleküler yöntemlerle inceleme sonuçlarının değerlendirilmesi amaçlanmıştır. Ülkemizde yerli olguların görülme sıklığı düşük olmasına rağmen son yıllarda artan göçmen sayısı nedeniyle leishmaniasis tüm dünyada olduğu gibi ülkemizde de yeniden önem kazanmıştır. Visseral leishmaniasis tanısında özellikle kemik iliği ve buffy coat örnekleri; kütanöz leishmaniasis tanısında ise yara aspiratı veya doku biyopsi örneklerinden yararlanılmaktadır. Kemik iliği ve doku biyopsi örneklerinin alınması hasta açısından oldukça zahmetli olmakta, ayrıca doğru şekilde alınmadığında yanlış tanıya yol açabilmektedir. Bu nedenle mümkün olduğunca birden fazla yöntemin bir arada kullanılması ile tanı koyma şansının artırılması önerilmektedir.Yöntem: Ocak 2015 ile Temmuz 2018 tarihleri arasında leishmaniasis şüphesiyle Halk Sağlığı Genel Müdürlüğü Ulusal Parazitoloji Referans Laboratuvarı’na gönderilen 271 adet kan, kemik iliği, doku ve/veya yara aspiratı örneğinin NNN besiyerine ekimi yapılmış ve ticari kit kullanılarak Genesig, Primer Design, UK realDesign, UK for real-time PCR method. Results: While 45 16.60% of the samples were evaluated as positive by any method, 226 83.40% were evaluated as negative. Of the 174 64.20% samples evaluated by PCR alone, 22 12.64% were positive and 152 87.36% were negative. Of the 52 19.20% samples evaluated by culture method only, 7 13.46% were positive and 45 86.54% were negative. And finally, of the 45 16.60% samples evaluated by both methods, 29 64.45% were negative by both PCR and culture, 10 22.22% were positive by PCR and culture, 6 13.33% were positive by PCR and negative by culture. There were no samples detected negative by PCR and positive by culture. There were no samples detected negative by PCR and positive by culture. A moderate concordance was found between the two methods studied κ= 0.545 . Conclusion: Since the sensitivity and specificity of the two diagnostic methods are different, it is concluded that the use of a combination of culture and PCR methods that allow the growth of the parasite by culture, especially in the case of low parasitemia and amplification of DNA by PCR, will increase the chance of diagnosis of the disease

Keywords

Leishmaniasis, kültür, NNN besiyeri, real-time PCR

Evaluation of culture and PCR results of leishmaniasis suspected samples

Abstract

Objective: In this study, we aimed to evaluate the results of blood, bone marrow and / or tissue biopsy and wound aspirate samples sent to our laboratory with suspicion of leishmaniasis on Novy-MacNeal-Nicolle NNN medium and the results of molecular examination. Although incidence of indigenous cases in our country is low, due to increasing number of migrants in recent years, leishmaniasis has regained importance in our country as in the whole world. Especially bone marrow and buffy coat samples for the diagnosis of visceral leishmaniasis and wound aspirate or tissue biopsy specimens for the diagnosis of cutaneous leishmaniasis are used. Obtaining bone marrow and tissue biopsy specimens is very painful for the patient and may lead to misdiagnosis if not obtained correctly. Therefore, it is recommended to increase the chance of diagnosis by using more than one method together.Methods: Two hundred seventy one blood, bone marrow, tissue and / or wound aspirate specimens between January 2015 and July 2018 were sent to the National Parasitology Reference Laboratory of the General Directorate of Public Health on suspicion of leishmaniasis and cultured in NNN medium were then processed by using a commercial kit Genesig, Primer Design, UK for real-time PCR method. Results: While 45 16.60% of the samples were evaluated as positive by any method, 226 83.40% were evaluated as negative. Of the 174 64.20% samples evaluated by PCR alone, 22 12.64% were positive and 152 87.36% were negative. Of the 52 19.20% samples evaluated by culture method only, 7 13.46% were positive and 45 86.54% were negative. And finally, of the 45 16.60% samples evaluated by both methods, 29 64.45% were negative by both PCR and culture, 10 22.22% were positive by PCR and culture, 6 13.33% were positive by PCR and negative by culture. There were no samples detected negative by PCR and positive by culture. There were no samples detected negative by PCR and positive by culture. A moderate concordance was found between the two methods studied κ= 0.545 . Conclusion: Since the sensitivity and specificity of the two diagnostic methods are different, it is concluded that the use of a combination of culture and PCR methods that allow the growth of the parasite by culture, especially in the case of low parasitemia and amplification of DNA by PCR, will increase the chance of diagnosis of the disease

Keywords

Leishmaniasis, culture, NNN medium, real-time PCR

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