COMPARISON OF VALIDITY OF FNACAND ZIEHL–NEELSEN STAINS IN DIAGNOSING CERVICAL TUBERCULOUS LYMPHADENITIS

Objective: To compare the validity of FNAC & ZN staining in diagnosing tuberculous lymphadenitis taking mycobacterial culture as gold standard. Study Design: Cross sectional study. Place and Duration of Study: Pathology department, Fauji Foundation Hospital Peshawar, from Oct 2019 to Sep 2020. Methodology: This study includes 100 patients with enlarged cervical lymph nodes who presented as outdoor patients. After routine baseline & radiological investigations, FNAC & ZN staining was carried out, followed by mycobacterial culture. Results: FNAC of tuberculous patients was positive in 56 (56%) patients while 24 (24%) patients revealed posi-tive mycobacterial culture. Inpatients with positive culture, 77 (77%) were FNAC positive and 23 (23%) were negative on cytology. Conclusion: Fine needle aspiration is more sensitive & specific test than Ziehl-Neelsen staining.

Retrospective diagnosis of lymphatic tuberculosis in frozen samples using two genetic amplification methods, Xpert® MTB/RIF ULTRA and Abbott RealTime MTB Assay

Objectives. The main objective of the present study is to assess the sensitivity and specificity of a retrospective diagnostic of lymphatic tuberculosis (LTB), testing frozen samples using gene amplification PCR methods. The secondary objective was to compare the results of two different commercial tuberculosis gene amplification methods for this purpose. Material and methods. We retrospectively studied 38 frozen samples, previously processed for mycobacterial culture between January 2014 and August 2019. The results of the previous cultures were: 21 samples positive for Mycobacterium tuberculosis complex (MTB) (5 being smear positive), 7 samples culture positive for Mycobacterium avium-intracellulare complex and 10 samples which were mycobacterial culture negative and discarded for LTB diagnosis, used as controls. The samples were processed using two gene amplification methods: Xpert® MTB/RIF Ultra (Cepheid) and Abbott RealTime MTB Assay (Abbott). Results. Compared to initial culture results the sensitivity and specificity of Xpert® MTB/RIF Ultra were 57.1% and 100% and 52.3 % and 92.5%, respectively for the Abbott RealTime MTB assay. The differences were not statiscally significant. In addition, there were no differences according to the period of freezing. Conclusions. Gene amplification of frozen samples confirmed the diagnosis of lymphatic TB in almost 60% of cases, allowing retrospective diagnosis in initially non suspected cases. Both gene amplification techniques tested were equally useful.

Outcomes of adjunctive surgery for nontuberculous mycobacterial pulmonary disease

Background Owing to the unsatisfactory results of antibiotic treatment alone, surgical resection is currently considered as adjunctive therapy in patients with nontuberculous mycobacterial pulmonary disease (NTM-PD). However, reports regarding the outcomes of surgery vary considerably by institution. Here, we investigated the surgical outcomes and risk factors associated with unfavorable outcomes after surgery. Methods We analyzed patients with NTM-PD who underwent pulmonary resection at Seoul National University Hospital between January 1, 2006, and December 31, 2020, and assessed the types of surgical procedures, complications, and long-term outcomes. Multivariate logistic regression analysis was used to identify the risk factors associated with treatment refractoriness or recurrence after surgery. Results Among 67 patients who underwent surgery during the study period, the most common indication for surgery was persistent culture positivity despite rigorous medical treatment (80.6%), followed by longstanding cavitary lesions or radiographic aggravation (10.4%) and massive hemoptysis (4.5%). Among 53 patients with positive mycobacterial cultures at the time of surgery, 38 (71.7%) achieved initial negative culture conversion, 9 (17.0%) of whom experienced recurrence. Nine (13.4%) patients experienced postoperative complications, which were managed without lasting morbidity and mortality. Female sex (adjusted odds ratio [aOR] 6.63; 95% confidence interval [CI] 1.04–42.4; P = .046), preoperative positive mycobacterial culture (aOR 5.87; 95 %CI 1.04–33.08; P = .045), and residual lesions (aOR 6.86; 95 %CI 1.49–31.56; P = .013) were associated with refractoriness or recurrence. Conclusions Pulmonary resection is a reasonable treatment modality for patients with refractory NTM-PD or major complications such as massive hemoptysis. The potential risk factors associated with unfavorable outcomes included female sex, preoperative positive mycobacterial culture, and residual lesions after surgery.

Improved detection of Mycobacterium tuberculosis and M. bovis in African wildlife samples using cationic peptide decontamination and mycobacterial culture supplementation

In South Africa, mycobacterial culture is regarded as the gold standard for the detection of Mycobacterium tuberculosis complex (MTBC) infection in wildlife even though it is regarded as “imperfect.” We compared a novel decontamination and mycobacterial culture technique (TiKa) to the conventional mycobacterium growth indicator tube (MGIT) system using known amounts of bacilli and clinical samples from MTBC-infected African buffaloes ( Syncerus caffer), white rhinoceros ( Ceratotherium simum), and African elephants ( Loxodonta africana). Use of the TiKa-KiC decontamination agent on samples spiked with 10,000 to 10 colony forming units (cfu) of M. bovis (SB0121) and M. tuberculosis (H37Rv) had no effect on isolate recovery in culture. In contrast, decontamination with MGIT MycoPrep resulted in no growth of M. bovis samples at concentrations < 1,000 cfu and M. tuberculosis samples < 100 cfu. Subsequently, we used the TiKa system with stored clinical samples (various lymphatic tissues) collected from wildlife and paucibacillary bronchoalveolar lavage fluid, trunk washes, and endotracheal tube washes from 3 species with known MTBC infections. Overall, MTBC recovery by culture was improved significantly ( p < 0.01) by using TiKa compared to conventional MGIT, with 54 of 57 positive specimens versus 25 of 57 positive specimens, respectively. The TiKa mycobacterial growth system appears to significantly enhance the recovery of MTBC members from tissue and paucibacillary respiratory samples collected from African buffaloes, African elephants, and white rhinoceros. Moreover, the TiKa system may improve success of MTBC culture from various sample types previously deemed unculturable from other species.

Outcomes of Adjunctive Surgery for Nontuberculous Mycobacterial Pulmonary Disease

Background Owing to the unsatisfactory results of antibiotic treatment alone, surgical resection is currently considered as adjunctive therapy in patients with nontuberculous mycobacterial pulmonary disease (NTM-PD). However, reports regarding the outcomes of surgery vary considerably by institution. Here, we investigated the surgical outcomes and risk factors associated with unfavorable outcomes after surgery. Methods We analyzed patients with NTM-PD who underwent pulmonary resection at Seoul National University Hospital between January 1, 2006, and December 31, 2020, and assessed the types of surgical procedures, complications, and long-term outcomes. Multivariate logistic regression analysis was used to identify the risk factors associated with treatment refractoriness or recurrence after surgery. Results Among 67 patients who underwent surgery during the study period, the most common indication for surgery was persistent culture positivity despite rigorous medical treatment (80.6%), followed by longstanding cavitary lesions or radiographic aggravation (10.4%) and massive hemoptysis (4.5%). Among 53 patients with positive mycobacterial cultures at the time of surgery, 38 (71.7%) achieved initial negative culture conversion, 9 (17.0%) of whom experienced recurrence. Nine (14.3%) patients experienced postoperative complications, which were managed without lasting morbidity and mortality. Female sex (adjusted odds ratio [aOR], 6.63; 95% confidence interval [CI], 1.04–42.4; P = .046), preoperative positive mycobacterial culture (aOR, 5.87; 95%CI, 1.04–33.08; P = .045), and residual lesions (aOR, 6.86; 95%CI, 1.49–31.56; P = .013) were associated with refractoriness or recurrence. Conclusions Pulmonary resection is a reasonable treatment modality for patients with refractory NTM-PD or major complications such as massive hemoptysis. The potential risk factors for unfavorable outcomes included female sex, preoperative positive mycobacterial culture, and residual lesions after surgery.

Detection of differentially culturable tubercle bacteria in sputum using mycobacterial culture filtrates

Rapid detection of tuberculosis (TB) infection is paramount to curb further transmission. The gold standard for this remains mycobacterial culture, however emerging evidence confirms the presence of differentially culturable tubercle bacteria (DCTB) in clinical specimens. These bacteria do not grow under standard culture conditions and require the presence of culture filtrate (CF), from axenic cultures of Mycobacterium tuberculosis (Mtb), to emerge. It has been hypothesized that molecules such as resuscitation promoting factors (Rpfs), fatty acids and cyclic-AMP (cAMP) present in CF are responsible for the growth stimulatory activity. Herein, we tested the ability of CF from the non-pathogenic bacterium Mycobacterium smegmatis (Msm) to stimulate the growth of DCTB, as this organism provides a more tractable source of CF. We also interrogated the role of Mtb Rpfs in stimulation of DCTB by creating recombinant strains of Msm that express Mtb rpf genes in various combinations. CF derived from this panel of strains was tested on sputum from individuals with drug susceptible TB prior to treatment. CF from wild type Msm did not enable detection of DCTB in a manner akin to Mtb CF preparations and whilst the addition of RpfABMtb and RpfABCDEMtb to an Msm mutant devoid of its native rpfs did improve detection of DCTB compared to the no CF control, it was not statistically different to the empty vector control. To further investigate the role of Rpfs, we compared the growth stimulatory activity of CF from Mtb, with and without Rpfs and found these to be equivalent. Next, we tested chemically diverse fatty acids and cAMP for growth stimulation and whilst some selective stimulatory effect was observed, this was not significantly higher than the media control and not comparable to CF. Together, these data indicate that the growth stimulatory effect observed with Mtb CF is most likely the result of a combination of factors. Future work aimed at identifying the nature of these growth stimulatory molecules may facilitate improvement of culture-based diagnostics for TB.

Bronchial brushing Xpert improves the diagnostic efficiency of sputum Xpert in patients with pulmonary tuberculosis

Background: The role of bronchial brushing Xpert MTB/RIF (hereafter referred to as Xpert) in patients with pulmonary tuberculosis (PTB) remains unclear. Therefore, a retrospective study was conducted aiming to evaluate the diagnostic efficiency of bronchial brushing Xpert in patients with PTB. Methods: Between July 2018 and August 2019, suspected PTB patients who were admitted to our hospital and had bronchial brushing and matched sputum collection for acid-fast bacilli (AFB) smear, mycobacterial culture, and Xpert were included for further analysis. Subsequently, PTB was defined based on mycobacterial culture, and if an alternative diagnosis was established, ‘non-tuberculosis (TB)’ was considered. Comparison of bronchial brushing and matched sputum examination was performed between groups. Then, the differences in the sensitivities between bronchial brushing and sputum Xpert were examined using the chi-square test. Results: A total of 111 patients were included and divided into TB and non-TB groups (52 versus 59 patients). The sensitivities of Xpert against culture were calculated as follows: sputum, 44.2% (23/52); bronchial brushing, 59.6% (31/52); sputum and bronchial brushing, 69.2% (36/52). The specificities of all Xpert assays were the same (100.0%, 59/59). A significant difference was found in the comparison of the sensitivities of Xpert using sputum, bronchial brushing and both, and the sensitivity of Xpert on both sputum and bronchial brushing was higher than that on sputum alone ( p < 0.05). Moreover, it appears that bronchial brushing Xpert was more sensitive than sputum Xpert in the detection of PTB. Conclusion: Bronchial brushing Xpert improves the diagnostic efficiency of sputum Xpert in the detection of PTB.