ABSTRACT
A total of 312 sputum samples from pediatric patients presumptive of multidrug resistant tuberculosis were tested for the detection of drug resistance using the GenoTypeMTBDRplus assay. A total of 193 [61.8%] patients were smear positive and 119 [38.1%] were smear negative by Ziehl-Neelsen staining. Line probe assay [LPA] was performed for 208 samples/cultures [193 smear positive samples and 15 cultures from smear negative samples]. Valid results were obtained from 198 tests. Of these, 125/198 [63.1%] were sensitive to both rifampicin [RIF] and isoniazid [INH]. 73/198 [36.9%] were resistant to at least INH/RIF, out of which 49 [24.7%] were resistant to both INH and RIF [multidrug resistant]. Children with tuberculosis are often infected by someone close to them, so strengthening of contact tracing in the program may help in early diagnosis to identify additional cases within the household. There is a need to evaluate newer diagnostic assays which have a high sensitivity in the case of smear negative samples, additional samples other than sputum among young children not able to expectorate, and also to fill the gap between estimated and reported cases under the program
ABSTRACT
Tuberculosis [TB] is caused by Mycobacterium tuberculosis [MTB] and the disease has remained a major health problem in most of the developing countries, particularly after the emergence of multidrug-resistant TB [MDR-TB]. The MDR-TB is an intriguing subject and very little is known about the in vivo processes which take place during the acquisition of MDR. This study describes a unique case of pulmonary TB [PTB] from which four sequential isolates of MTB could be isolated while the patient was on anti-tubercular treatment. The first baseline isolate was sensitive to all drugs, but the subsequent three isolates acquired resistance to multiple drugs and finally the patient died after 27 months post-diagnosis when his fourth isolate became resistant to isoniazid, rifampicin, ethambutol and kanamycin. All sequential cultures were identified as MTB using conventional and molecular methods, including 16s RNA sequencing and the spoligotyping. Spoligotyping followed by comparison with SITVITWEB database revealed that all the isolates belonged to the family of the Central Asian Strain Delhi [CAS1_Delhi, ST26] genotype, and no cross or mixed infections were observed. The drug resistance was further characterized at the molecular level by sequencing the target genes [katG, inhA, rpoB, embB, eis promoter region and rrs]. The results revealed mutated alleles associated with resistance to the respective drugs. This unique case indicates that it is possible to isolate MTB during treatment if the strain is acquiring resistance. The data presented from four sequential isolates provides an insight into what sequential genetic and proteomic changes occur in the bacteria during the in vivo acquisition of MDR.
Subject(s)
Humans , Male , Mycobacterium tuberculosis , GenotypeABSTRACT
Collection and processing of sputum samples for the detection of acid fast bacilli [AFB] is hazardous for health-workers in developing countries with limited facilities. The phenol ammonium sulfate [PhAS] method involves smear microscopy and Ziehl-Neelson [ZN] staining of precipitates/ floccules formed in sputum samples when PhAS is added. The present study has been designed to assess the performance and safety of this method. The study was conducted from January 2011 to March 2011 at the Department of Microbiology, Lala Ram Sarup Institute of Tuberculosis and Respiratory Diseases, New Delhi. A total of 1038 sputum samples were subjected to ZN staining before and after treatment with PhAS. The smear microscopy results of the PhAS treated and untreated samples were compared. In addition, 200 representative samples were inoculated after processing by petroff's method directly for culture and after treatment with PhAS. The sensitivity, specificity, positive predictive value and negative predictive value of the PhAS solution treated ZN smear microscopy method were found to be 98.8%, 88.5%, 98.0% and 92.7% respectively in comparison with direct smear microscopy. The overall correlation between the two methods was found to be 97.3%. None of the PhAS treated samples grew Mycobacterium tuberculosis on culture. Sputum microscopy with PhAS solution is a safe, reliable and inexpensive alternative for direct microscopy. This method can be conveniently applied for usage in microscopy centers with limited bio-safety facilities