Systematic assessment of clinical and bacteriological markers for tuberculosis reveals discordance and inaccuracy of symptom-based diagnosis for treatment response monitoring

This work was supported by Commonwealth PhD studentship award to Dr Bariki Mtafya (Award number: TZS-2016-718) at University of St Andrews and European and Developing Countries Clinical Trials Partnership through TWENDE project (grant number; TWENDE-EDCTP-CSA-2014-283) and PanACEA II (grant number; 97118-PanACEA-TRIA.2015.1102) awarded to Professor Stephen Gillespie and Dr Wilber Sabiiti at the University of St Andrews, UK.Background : Clinical symptoms are the benchmark of tuberculosis (TB) diagnosis and monitoring of treatment response but is not clear how they relate to TB bacteriology, particularly the novel tuberculosis Molecular Bacterial Load Assay (TB-MBLA). Methods : Presumptive cases were bacteriologically confirmed for TB and assessed for symptom and bacteriological resolution using smear microscopy (SM), culture and TB-MBLA over 6-month treatment course. Kaplan Meier and Kappa statistics were used to test relationship between symptom- and bacteriological-positivity. Results : A cohort of 46 bacteriologically confirmed TB cases were analysed for treatment response over a six-month treatment course. Pre-treatment symptom and bacteriological positivity concurred in over 70% of the cases. This agreement was lost in over 50% of cases whose chest pain, night sweat, and loss of appetite had resolved by week 2 of treatment. Cough resolved at a 3.2% rate weekly and was 0.3% slower than the combined bacteriological (average of MGIT and TB-MBLA positivity) resolution rate, 3.5% per week. Drop in TB-MBLA positivity reflected fall in bacillary load, 5.7±1.3- at baseline to 0.30±1.0- log10 eCFU/mL at month 6, and closer to cough resolution than other bacteriological measures, accounting for the only one bacteriologically positive case out of seven still coughing at month 6. Low baseline bacillary load patients were more likely to be bacteriologically negative, HR 5.6, p=0.003 and, HR 3.2, p=0.014 by month-2 and 6 of treatment respectively. Conclusion : The probability of clinical symptoms reflecting bacteriological positivity weakens as patient progresses on anti-TB therapy, making symptom-based diagnosis a less reliable marker of treatment response.Publisher PDFPeer reviewe

Extracting and classifying salient fields of view from microscopy slides of tuberculosis bacteria

Funding: We will like to express our appreciation to the McKenzie Institute for providing the necessary funding to complete this work.Tuberculosis is one of the most serious infectious diseases, and its treatment is highly dependent on early detection. Microscopy-based analysis of sputum images for bacilli identification is a common technique used for both diagnosis and treatment monitoring. However, it a challenging process since sputum analysis requires time and highly trained experts to avoid potentially fatal mistakes. Capturing fields of view (FOVs) from high resolution whole slide images is a laborious procedure, since they are manually localized and then examined to determine the presence of bacteria. In the present paper we propose a method that automates the process, thus greatly reducing the amount of human labour. In particular, we (i) describe an image processing based method for the extraction of a FOV representation which emphasises salient, bacterial content, while suppressing confounding visual information , and(ii) introduce a novel deep learning based architecture which learns from coarsely labelled FOV images and the corresponding binary masks, and then classifies novel FOV images as salient (bacteria containing) or not. Using a real-world data corpus, the proposed method is shown to out-perform 12 state of the art methods in the literature, achieving (i) an approximately 10% lower overall error rate than the next best model and(ii) perfect sensitivity (7% higher than the next best model).PostprintPostprin

A practical approach to render tuberculosis samples safe for application of tuberculosis Molecular Bacterial Load Assay in clinical settings without a biosafety level 3 laboratory

Funding: This work was supported by the United Kingdom Research and Innovation (UKRI) for the TB-MBLA translation study in routine healthcare settings through the Food Security and Health for East Africa Project (Grant number# EP/T01525X/1) as part of postdoctoral work for Dr Bariki Mtafya at the National Institute for Medical Research Centre-Mbeya Medical Research Centre, Mbeya, Tanzania with the University of St Andrews, UK.Background Mycobacterium tuberculosis is a category B pathogen requiring level-3-containment laboratories for handling. We assessed the efficacy of heat and Guanidine thiocyanate (GTC) to inactivate M. tuberculosis prior to performance of tuberculosis Molecular Bactrial Load Assay (TB-MBLA). Method We performed in vitro experiments using H37Rv reference strain and replicated in sputum specimens. A 0.5 MacFarland standard of M. tuberculosis was serially diluted to 1x10 CFU/mL and pooled sputum was homogenised prior to serial dilutions and Xpert MTB/RIF Ultra. Three replicates for each containing 1 mL for M. tuberculosis and sputum were inactivated at 80 °C for 20 minutes and with GTC for 15 minutes. Inactivated samples were processed for culture and TB-MBLA. Results No M. tuberculosis growth was observed in MGIT for GTC or heat treated H37Rv cultures. All untreated H37Rv dilutions were MGIT positive except the most diluted specimens. Heat and GTC treatment of H37Rv reduced TB-MBLA load by 2.1log10 (P = 0.7) and 1.8log10 (P = 0.7) respectively, compared to controls. In contrast, heat treated sputum had TB-MBLA bacterial load of 3.47 ± 3.53 log10 compared to 5.4 ± 3.1 log10 eCFU/mL for GTC (p = 0.57). All heat and GTC treated sputum were culture negative. Conclusion Heat or GTC renders M. tuberculosis non-viable and eliminates the need for BSL3 laboratory for performing TB-MBLA in routine healthcare settings.Publisher PDFPeer reviewe