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AMAÇ: Tüberküloz (TB), Mycobacterium tuberculosis kompleks (MTBK) grubuna ait mikroorganizmaların neden olduğu kronik gidişatlı, bir enfeksiyon hastalığıdır. Anti tüberküloz ilaçlar ile tedavi edilmekte ve kontrol altına alınmakta olan bu hastalık; bazen ilaçlara yanıt vermeyebilir. Tüberkülozda ilaç direnç araştırması yapılması tüberküloz kontrol ve tedavi programlarının temelini oluşturmaktadır.GEREÇ VE YÖNTEM: Çalışmamızda, laboratuvarımızda izole edilen MTBK izolatlarının primer anti tüberküloz ilaçlara duyarlılıklarının araştırılması ve direnç paternlerinin belirlenmesi amaçlanmıştır. Kliniğimizde 2011 - 2019 yılları arasında tüberküloz şüphesi ile mikrobiyoloji laboratuvarına gönderilen 8804 klinik örnek değerlendirmeye alınmıştır.BULGULAR: Çalışmaya alınan örneklerden 126’sında tüberküloz kültürü pozitif gelmiştir. 126 örneğe (%1.4) anti-tüberküloz duyarlılık testi uygulanmıştır. 28 (%22.3) izolat ise test edilen anti tüberküloz ilaçlardan en az birine dirençli olarak tespit edilmiştir. Çok ilaca direnç ise %2,38 oranında bulunmuştur.SONUÇ: Tüberküloz tedavisinde direnç gelişimini önlemek ve tedavinin sağlıklı bir şekilde yapılabilmesi için her hastaya kültür ve antibiyotik duyarlılık testlerinin çalışılması, verilerin elde edilmesi gerekmektedir., OBJECTIVE: Tuberculosis (TB) is a chronic infectious disease caused by microorganisms belonging to the Mycobacterium tuberculosis complex (MTBK) group. This disease is treated and controlled with antituberculosis drugs; but sometimes it may not respond to medications. Investigation of drug resistance in tuberculosis, forms the basis of tuberculosis control and treatment programs.MATERIAL AND METHODS: In our study, it was aimed to investigate the susceptibility of MTBK isolate, isolated in our laboratory to primary antituberculosis drugs and to determine their resistance patterns. Between 2011 and 2019, 8804 clinical samples sent to the microbiology laboratory with the suspicion of tuberculosis were evaluated in our clinic.RESULTS: Tuberculosis culture was positive in 126 of the samples included in the study. Anti-tuberculosis susceptibility test was applied to 126 samples (1.4%). 28 (22.3%) isolates were found to be resistant to at least one of the tested antituberculosis drugs tested. Multidrug resistance was found at the rate of 2.38%.CONCLUSIONS: In order to prevent the development of resistance in the treatment of tuberculosis and to provide a healthy treatment, culture and antibiotic sensitivity tests should be performed for each patient and data should be obtained.

Ceyhan, İsmail (Balikesir Author), Members of the Mycobacterium tuberculosis complex (MTBC) are the causative agents of tuberculosis (TB). Rifampin resistance in the clinical isolates of MTBC is an important indicator for multidrug resistant-TB (MDR-TB) cases. In this study, it was aimed to evaluate whether the Sensititre MycoTB plaque method is suitable for the routine use in determining drug susceptibility of rifampin resistant MTBC strains. Xpert MTBC/RIF positive rifampin resistant 100 MTBC isolates were included in the study. Xpert MTBC/RIF (Cepheid, USA) test were performed after the samples were processed by homogenization and decontamination and acid-fast staining. Rifampin resistant clinical samples were cultured in automated MGIT/BACTEC 960 (Becton Dickinson, USA) system and acid fast bacteria (AFB) were investigated. The anti-TB drug susceptibility tests of all culture positive AFB and cord factor identified as MTBC by using a cart test (MPB64, Capilla TB-Neo, Tauns Laboratories, Inc., Numazu, Japan) were performed with the Lowenstein-Jensen proportion method (LJPM) and Sensititre MycoTB (Trek Diagnostic Systems, Cleveland, OH, USA) methods. For the comparison of the methods used, the tests were performed simultaneously. The standard LJPM was performed according to the previously described procedures by World Health Organization and the Sensititre MycoTB plate method was performed as defined by the manufacturer. The final concentrations of isoniazide, rifampin, rifabutin, ethambutol, ofloxacin, moxifloxacin amikacin, kanamycin, cycloserine, ethionamide and p-aminosalicylic acid in Lowenstein-Jensen media for LJPM were 0.2 mu g/ml, 40.0 mu g/ml, 20.0 mu g/ml, 2.0 mu g/ml, 2.0 mu g/ml, 1.0 mu g/ml, 4.0 mu g/ml, 30.0 mu g/ml, 30.0 mu g/ml, 40.0 mu g/ml, 40.0 mu g/ml and 1.0 mu g/ml, respectively. The results were obtained in 14 days for all of the drugs in the Sensititre MycoTB plate method and in 28-42 days in the LJPM. In this study, the sensitivity and specificity percentages of the Sensititre MycoTB method and the categorical agreement between the two methods were calculated. The sensitivity and specificity percentages of the Sensititre MycoTB plate method were between 84.4-100% and 95.6-6100%, respectively. The categorical agreements between the two methods were 92-100% for the drugs tested in the study. Ethambutol was found to have the lowest sensitivity (84.4%) and specificity (95.6%). The sensitivities of isoniazide, ofloxacin, streptomycin, kanamycin, ethionamide and p-aminosalicylic acid were 98.8%, 90.0%, 94.3%, 87.5%, 91.7% and 95.6%, respectively, while rifampicin, rifabutin, moxifloxacin, amikacin, cycloserine were calculated as 100%. The specificities of isoniazid, rifampicin, rifabutin, ofloxacin, moxifloxacin, amikacin, kanamycin, and cycloserine were found to be 100%, streptomycin, ethionamide and p-aminosalicylic specificity were 96.9%, 97.4% and 98.9% respectively. The categorical agreement was 96-100% in all tested drugs except ethambutol (92%). As a result, although the cost is high, owing to the short incubation period, easy to perform, the possibility for evaluating both first and second line anti-TB drugs simultaneously, determination of minimum inhibitory concentration values of the drugs, long shelf life, high sensitivity, specificity and the categorical agreement values, the Sensititre MycoTB method was determined as an effective method that can be used especially in laboratories where the workload and the MDR-TB cases are high., Mycobacterium tuberculosis kompleks (MTBK) üyeleri, tüberküloz (TB) hastalığı etkenidir. Klinik MTBK izolatlarında saptanan rifampisin direnci, çok ilaca dirençli tüberküloz (ÇİD-TB) olgularının önemli bir göstergesidir. Bu çalışmada, rifampisine dirençli MTBK izolatlarının ilaç duyarlılığını belirlemede Sensititre MycoTB plak yönteminin, rutin kullanıma uygun olup olmadığının değerlendirilmesi amaçlanmıştır. Çalışmaya, Xpert MTBC/RIF (Cepheid, ABD) test sonucu pozitif rifampisine dirençli 100 MTBK izolatı dahil edilmiştir. Hasta örnekleri, homojenizasyon ve dekontaminasyon işlemi uygulandıktan sonra aside dirençli boyamanın ardından Xpert MTBC/RIF testine alınmıştır. Rifampisine dirençli klinik örnekler, MGIT BACTEC 960 (Becton Dickinson, ABD) sisteminde kültürü yapılarak aside dirençli bakteri (ARB) araştırılmıştır. Kültürden izole edilen ARB ve kord faktör pozitif bakterilere kart testi (MPB64, Capilla TB-Neo, Tauns Laboratories, Inc., Numazu, Japonya) uygulanmış ve MTBK olarak saptanan izolatların Löwenstein-Jensen proporsiyon yöntemi (LJPY) ve Sensititre MycoTB plak (Trek Diagnostic Systems, Cleveland, OH, ABD) yöntemiyle anti-TB ilaç duyarlılık testleri çalışılmıştır. Yöntemler arasında karşılaştırma yapmak amacıyla testler aynı anda çalışılmıştır. LJPY, Dünya Sağlık Örgütü tarafından daha önce tarif edildiği gibi, Sensititre MycoTB testi ise üretici firmanın tanımladığı şekilde çalışılmış ve değerlendirilmiştir. LJPY’de izoniazid, rifampisin, rifabutin, etambutol, ofloksasin, moksifloksasin, streptomisin, amikasin, kanamisin, sikloserin, etionamid ve p-aminosalisilik asit için final konsantrasyonlar sırasıyla; 0.2 µg/ml, 40.0 µg/ml, 20.0 µg/ml, 2.0 µg/ml, 2.0 µg/ml, 1.0 µg/mL, 4.0 µg/ml, 30.0 µg/ml, 30.0 µg/ml, 40.0 µg/ml, 40.0 µg/ml ve 1.0 µg/ml olacak şekilde hazırlanmıştır. Sensititre MycoTB plak yönteminde tüm ilaçlar için 14. günde, LJPY’de ise 28. ve 42. günlerde sonuç alınmıştır. Çalışmada, Sensititre MycoTB testinin duyarlılık ve özgüllük oranları ile iki test arasındaki kategorik uyum hesaplanmıştır. Sensititre MycoTB plak yönteminin duyarlılığı ve özgüllüğünün sırasıyla %84.4-100 ve %95.6-100 aralığında olduğu bulunmuştur. Bu çalışmada test edilen ilaçlar için iki yöntem arasındaki kategorik uyum %92-100 olarak saptanmıştır. Etambutolün en düşük duyarlılık (%84.4) ve özgüllük (%95.6) değerlerine sahip ilaç olduğu tespit edilmiştir. İzoniazid, ofloksasin, streptomisin, kanamisin, etionamid ve p-aminosalisilik asit duyarlılıklarının sırasıyla %98.8, %90.0, %94.3, %87.5, %91.7 ve %95.6; rifampisin, rifabutin, moksifloksasin, amikasin, sikloserin duyarlılıklarının ise hepsinde %100 olduğu saptanmıştır. İzoniazid, rifampisin, rifabutin, ofloksasin, moksifloksasin, amikasin, kanamisin ve sikloserinin özgüllükleri %100, streptomisin, etionamid ve p-aminosalisilik asidin özgüllükleri ise sırasıyla %96.9, %97.4 ve %98.9 olarak bulunmuştur. Test edilen tüm ilaçlarda kategorik uyum, etambutol (%92) hariç %96-100 arasında saptanmıştır. Maliyetinin yüksek olmasına rağmen inkübasyon süresinin kısa olması, kolay uygulanabilmesi, birinci ve ikinci seçenek anti-TB ilaçların duyarlılıklarının aynı anda test edilebilmesi, ilaçların minimum inhibitör konsantrasyonu değerlerinin saptanabilmesi, raf ömrünün uzun olması, yüksek duyarlılık, özgüllük ve kategorik uyum oranları nedeniyle Sensititre MycoTB yönteminin, iş yükü ve ÇİD-TB olguları fazla laboratuvarlarda kullanılabilecek etkili bir yöntem olduğu sonucuna varılmıştır.

PubMed: 29933730, 2-s2.0-85054329350, Tuberculosis is still one of the most important public health problems worldwide. Due to the increase of multidrug-resistant (MDR) tuberculosis and extensively drug-resistant (XDR) tuberculosis cases, fast, practical and standardized methods need to be developed for the determination of drug resistance in Mycobacterium tuberculosis complex. The aim of this study was to compare the Sensititre MycoTB plate method with the gold standard agar proportion method (APM); in order to determine if the Sensititre MycoTB plate method is effective enough to be used instead of the APM. A total of 100 M.tuberculosis complex isolates, 36 with different resistance patterns and 64 sensitive to all first-line agents stored in culture collection of Ege University Faculty of Medicine Microbiology Department Laboratory, were used in the study. The susceptibility of rifampin, isoniazid, ethambutol, streptomycin, ofloxacin, moxifloxacin, amikacin, rifabutin, para-aminosalicylic acid, ethionamide, cycloserine and kanamycin for all the isolates were determined by APM and Sensititre MycoTB plate methods. APM was performed as defined by CLSI, Sensititre MycoTB plate method was performed as instructed by the manufacturer. Results were assessed in 95% confidence interval and sensitivity, specificity and categorical agreement values were determined. Results were obtained in 14 days using the Sensititre MycoTB plate method. Sensitivity and the specificity of the Sensititre MycoTB was between 86-100% and 95-100% respectively, while the categorical agreement between the two methods were 95-100% for the drugs tested. The sensitivities of the drugs were 100% except ethambutol and ethionamide. The specificity values for both of the drugs and categorical agreement were over 95%. All the drugs were in the 95% confidence interval (p< 0.001) and the greatest difference between the lower and upper levels of sensitivity and specificity confidence limits was found in ethambutol and ethionamide. MDR was detected in 16 isolates with the APM and in 20 isolates with the Sensititre MycoTB plate method. None of the isolates had XDR [in addition to MDR resistance against any fluoroquinolones and at Least one of the second-line parenteral drugs (amikacin, kanamycin or capreomycin)]. As a result, owing to the short Incubation period (14 days), high sensitivity, specificity and categorical agreement values, and the possibility for evaluating both first- and second-line agents and the quantitative values of drugs; the Sensititre MycoTB method was determined as an effective method that can be used especially in laboratories where the rate of MDR-TB isolates are high. © 2018 Ankara Microbiology Society. All rights reserved.

Mycobacterium tuberculosis kompleks' in neden olduğu tüberküloz hastalığı tanı ve tedavi yöntemlerindeki gelişmelere rağmen dünyada ve ülkemizde ciddi bir halk sağlığı sorunu olmaya devam etmektedir. Günümüzde, artan direnç nedeniyle etkili tedavi sağlanamaması, duyarlılık testleri ile bu direncin takibini gerekli hale getirmiştir. Son yıllarda moleküler yöntemlerdeki gelişmeler tüberküloz epidemiyolojisinin anlaşılmasını kolaylaştırmış, reenfeksiyon/reaktivasyon ayrımı, salgınların tanımlanması, hastalığın toplumdaki yayılımının ve laboratuvarlardaki çapraz kontaminasyonların takibinin sağlanması için yararlı olmuştur. Bu tez çalışmasında, 2013-2014 yıllarında Ege Üniversitesi mikobakteriyoloji laboratuvarında izole edilen M. tuberculosis kompleks izolatlarının genotiplendirilmesi, çok ilaca dirençli (ÇİD) izolatlarda ise ikinci seçenek tüberküloz ilaçlarından amikasin (AK) ve ofloksasin (OFL) duyarlılığının belirlenmesi, direnç ve genotip arasındaki ilişkinin değerlendirilmesi amaçlanmıştır. Yüzkırkiki izolatın spoligotiplendirilmesi ile edinilen verilere göre; T ailesi baskın bulunmuş, 2014 yılı izolatlarında ise bunu H ailesi (H1, H3, H4) takip etmiştir. MIRU-VNTR yöntemi ile tiplendirmek için seçilen, en az bir primer ilaca dirençli, 15 izolatın spoligotipleri değerlendirildiğinde beşi T1, ikisi LAM7TUR, ikisi Pekin, birer tanesi T2, U, H3, CAS1DELHI olarak tanımlanmış, iki tanesi ise veritabanındaki izolatların hiçbiriyle eşleşmemiştir. Spoligotiplerine göre Pekin, LAM7TUR ve CAS1DELHI ailesindeki suşlar MIRU-VNTR yöntemi ile de aynı gruplarda bulunmuştur. Ayrıca test edilen beş ÇİD izolatta AK ve OFL'ye karşı dirence rastlanmamıştır. Tüm dirençli kökenler değerlendirildiğinde bu çalışmada direnç ve genotipler arasında ilişki tespit edilememiştir. Sonuç olarak bu çalışmada direnç ve genotip arasında bir ilişki bulunmasa da tüberkülozun ve dirençli suşların yayılımının izlenmesi bakımından alternatif moleküler metotlar geliştirilmesi yönünde araştırmalara devam edilmeli ve bu yöntemlerin yaygın kullanımı sağlanmalıdır., Despite improvements in the diagnostic and therapeutic methods of tuberculosis caused by Mycobacterium tuberculosis complex, there still remains a serious public health problem in the world and in our country. Today, failure to provide effective treatment due to increased resistance has made it necessary to follow this resistance by susceptibility tests. In recent years, advances in molecular methods have facilitated the understanding of tuberculosis epidemiology and have helped to distinguish between re-infection / reactivation, identification of outbreaks, the spread of the disease in society, and follow-up cross-contamination in laboratories. In this thesis study, genotyping of M. tuberculosis complex isolates isolated in Ege University mycobacteriology laboratory in 2013-2014, determination of susceptibility of amikacin (AK) and ofloxacin (OFL) in second choice tuberculosis drugs in multidrug-resistant strains (MDR) were aimed. According to the data obtained by the spoligotyping of 142 isolates; T family was predominant, followed by H family (H1, H3, H4) in 2014 isolates. Five isolates were identified as T1, two as LAM7TUR, two as Beijing, one as T2, U, H3, CAS1DELHI, and two as none of the isolates in the database, when the spoligotype of 15 isolates selected for typing by the MIRU-VNTR method was evaluated. According to the spoligotype, Beijing, LAM7TUR and CAS1DELHI family strains were found in the same groups by the MIRU-VNTR method. In addition, resistance against AK and OFL was not detected in the five MDR isolates tested. When all resistant strains were evaluated, there was no relationship between resistance and genotypes in this study. In conclusion, although there is no relationship between resistance and genotype in this study, researches should be continued to develop alternative molecular methods in order to monitor the spread of tuberculosis and resistant strains and these methods should be widely used.

WOS: 000365510300023, PubMed ID: 26775391, Background/aim: The aim of this study is to assess the Xpert MTB/RIF assay for diagnosis of the Mycobacterium tuberculosis complex in clinical samples and to compare the results by reference to the diagnostic method, Bactec MGIT 960. Materials and methods: A total of 7407 samples were included from patients not primarily suggesting pulmonary or extrapulmonary tuberculosis (TB), collected from patients required to be screened for TB and excluding TB diagnoses since it was a differential diagnosis. Also included were a total of 411 samples from patients primarily suggesting pulmonary or extrapulmonary TB. Results: In the first group, 152 of 7407 samples yielded positive results with the Bactec MGIT 960, 131 (1.77%) were found positive with Lowenstein-Jensen medium, and 295 (3.99%) were found positive with Ziehl-Neelsen staining. In the second group, 24 (5.8%), 17 (4.1%), and 28 (6.8%) of 411 samples were found positive. Xpert MTB/RIF [27 (6.6%) of 411 samples] detected 3 additional samples as positive, and these 3 cases were clinically compatible with TB. Conclusion: The Xpert MTB/RIF assay shows superior performance for the diagnosis of TB. Its usefulness in culture-negative patients and the best method for integrating this diagnostic method into current tuberculosis diagnostic algorithms both need further study.

WOS: 000355774200014, PubMed ID: 26167828, It is well known that disseminated Mycobacterium bovis BCG infection is developed after BCG vaccination in infants with congenital cellular immune deficiencies such as mutations in genes along the interleukin (1)-12/interferon (IFN)-gamma pathway and mutations in nuclear factor-kB essential modulator (NEMO). In this report, a rifampicin-resistant M.bovis BCG strain isolated from an infant with NEMO defect was presented. An 8-month-old male infant with NEMO defect admitted to the pediatric outpatient clinic of our hospital with fever, generalized lymphadenopathy and hepatosplenomegaly. Microscopic examination of the smears, prepared from lymph node and liver biopsy specimens revealed abundant amount (3+) of acid-fast bacilli (AFB). Rifampicin-susceptible Mycobacterium tuberculosis complex (MTC) was detected by real-time PCR (GeneXpert MTB/RIF; Cepheid, USA) in the samples. The growth of mycobacteria was determined on the 20th day of culture performed in MGIT960 system (Becton Dickinson, USA). The isolate was identified as M.bovis BCG by GenoType MTBC kit (Hain Lifescience, Germany) and defined as M.bovis BCG [SIT 482 (BOV_1)] by spoligotyping. In the primary anti-tuberculosis drug susceptibility test performed by MGIT960 system, the isolate was found susceptible to rifampicin (RIF), isoniazid (INN), streptomycin (STM) and ethambutol (EMB). Then anti-tuberculosis treatment was started to the patient. However, the patient at the age of 2 years, re-admitted to the hospital with the complaint of hepatosplenomegaly. Smear of spontaneously draining abscess material obtained from subcutaneous nodules revealed intensive AFB positivity (3+) once again. In the present instance RIF-resistant MTC was detected with GeneXpert system in the specimen. The growth of mycobacteria was determined on the 13th day of culture and isolate was identified as M.bovis BCG. The present isolate was found susceptible to INN, STM and EMB but resistant to RIF. A mutation in the rpoB gene (codon 531, S531L) associated with RIF resistance was detected by using the partial sequencing of the rpoB gene. Patient died due to disseminated bovis BCG infection and multiple organ failure. To our knowledge, there are only six RIF-resistant M.bovis BCG strains isolated from patients in the literature. However, this is the first RIF-resistant M.bovis BCG strain isolated from a NEMO-deficient patient.

aim of this study was to identify the presence and prevalence of bovine tuberculosis in the eastern Anatolian border province of Van and its villages and to collect the preliminary data required for programs for the prevention and control of this zoonotic disease. The tuberculin test was performed on 210 cattle. Nasal swab and milk samples were obtained from the animals tested. Three cattle that had a positive tuberculin test also tested positive for the DNA target, a 580-bp fragment of IS6110 specific for members of the Mycobacterium tuberculosis complex. This fragment was also recovered from milk taken from the third animal tested positive for tuberculin. All the other cattle were negative for bovine tuberculosis on both tuberculin test and PCR assay. The results of the tuberculin test and the PCR were in close correlation with each other. Van is a border province in eastern Anatolia. In the city center and its villages, the prevalence of bovine tuberculosis was estimated as 1.42%. The aim of this study was to identify the presence and prevalence of bovine tuberculosis in the eastern Anatolian border province of Van and its villages and to collect the preliminary data required for programs for the prevention and control of this zoonotic disease. The tuberculin test was performed on 210 cattle. Nasal swab and milk samples were obtained from the animals tested. Three cattle that had a positive tuberculin test also tested positive for the DNA target, a 580-bp fragment of IS6110 specific for members of the Mycobacterium tuberculosis complex. This fragment was also recovered from milk taken from the third animal tested positive for tuberculin. All the other cattle were negative for bovine tuberculosis on both tuberculin test and PCR assay. The results of the tuberculin test and the PCR were in close correlation with each other. Van is a border province in eastern Anatolia. In the city center and its villages, the prevalence of bovine tuberculosis was estimated as 1.42%.