Yielding Accuracy of Morning and Spot Sputum Samples for Diagnosis Mycobacterium Tuberculosis Using Gene Xpert MTB/RIF Assay at EPHI, Addis Ababa, Ethiopia

Received: April 02, 2019 Revised: June 09, 2020 Accepted: June 20, 2020 Published Online: July 17, 2020 Background: Tuberculosis is a global health problem and leading cause of mortality and morbidity in the world. Correct diagnosis of tuberculosis is the main challenge for control of tuberculosis. World health organization has prioritized improving diagnostic guidelines and tests for TB diagnosis. Priority has been given for development of new Tuberculosis diagnostic methods. But diagnosis of Tuberculosis highly affected by quality specimen collected which has been given less attention. The aim of this is to assess Yielding accuracy of morning and spot sputum samples for diagnosis Mycobacterium tuberculosis using Gene Xpert Mycobacterium Tuberculosis (MTB)/ Rifampicin Resistance (RIF).


Introduction
Per year around 10.4 million people are affected globally by tuberculosis infection. Among these around 1.4 million are dead, with pulmonary type of TB is the main cause. The main challenge for global tuberculosis control is accurate and correct diagnosis of tuberculosis. World health organization has prioritized improving tuberculosis diagnosis guidelines and testing procedures. But the accuracy of laboratory diagnostic procedures affected by quality of sputum sample collection and processing. Nowadays the impact of quality sputum sample has been given less concern rather priority given for development of new TB diagnostic methods (Jochei & Akolhatkar, 1982;Charles et al., 2000). Most frequently used for laboratory diagnosis of TB is microscopy, which is inexpensive. But microscopy is less sensitive when the amount of acid fast bacilli low (most likely below 10,000 per ml not detected). Culture techniques for tuberculosis diagnosis have higher sensitivity than microscopic TB diagnosis. Gene Xpert MTB/RIF Assay has moderate sensitivity between culture techniques and microscopy. But Gene Xpert and culture techniques are more expensive than microscopy and only diagnosis when the amount of acid fast bacilli concentration is sufficient on the sputum sample (FMOH, 2012;Chemhuru et al., 2010;FMOH, 2014).
Inappropriate or poor quality sputum samples lead to false laboratory results for diagnosis of tuberculosis. Mostly poor sputum samples contain secretions from respiratory track with different amount saliva. The positivity of p.2 laboratory diagnosis of TB varies among different sputum samples from the same patient. For this reason more than one sputum sample is tested for suspected TB patients (Ethiopian Public Health Institute, the 2nd BSC Based EPHI's Strategic Management Plan, 2015/16 to 2019FMOH, 2014;Daniel, 2014;Minaleshewa et al., 2016).
Spot-morning-spot sputum samples collection methods were used for TB microscopy. Recommended sputum sample collection methods are different in different and their relative advantage for tuberculosis diagnosis is not well explained. Using spot sputum samples reduces possible transmission of tuberculosis. This study has been done to assess yielding of spot and morning sputum samples for tuberculosis diagnosis using Gene Xpert MTB/RIF Assay and possible effect of sputum sample quality on TB diagnosis.

Study Area
This study was conducted at Ethiopian Public Health Institute. It is located at Addis Ababa, capital city of Ethiopia. The institute is the center of health related researches, trainings and quality control of health service in Ethiopia.

Study Period and Study Design
Descriptive cross sectional study design was used to collect the data at Ethiopian Public Health Institute from April 1 to May 30, 2018 G.C.

Source population
All TB suspected patients living in Addis Ababa city and around the city were source population for the study.

Study population
All TB suspected patients requested to Ethiopian Public Health Institute TB national reference laboratory section during the study period.

Sample size and sampling technique
A total of two hundred twenty two (222) morning and spot sputum specimens from one hundred eleven (111) individual were analyzed using Gene Xpert MTB/RIF Assay. All suspected TB patients who came to EPHI for TB diagnosis during the study period were included.

Data Collection Procedure
Data was collected directly from M.TB suspected individuals by research participants and tested by Gene Xpert machine using both morning and spot sputum specimen. The sample was collected by clean, leak-proof and wide mouthed falcon tube about 2-4 ml of each morning and spot specimen. Gene Xpert MTB/RIF Assay used for testing the samples. If delay was unavoidable in testing, refrigeration 2-8 o c was used to store the specimen.
The whole process of patient preparation, specimen collection and testing were based on standard operating procedures manual. Mucoid and purulent sputum sample consider as good quality.

Data Processing and Analysis
The data was recorded, controlled and checked for its completeness everyday during the data collection period. All data was entered into an excel spreadsheet, and then transferred to SPSS version 20 for analysis. Descriptive statistics were performed and presented in tables and graph. Adjusted odds ratio and p value were used to assess relation of TB detection with sputum quality and consistency.

Ethical Consideration
Ethical clearance to do this research was obtained from Department of Medical Laboratory Science, Addis Ababa University. Written consent forms were agreed with patients to conduct the research.

Discussion
In this study, the detection of the morning and spot sputum samples were assesses using Gene Xpert MTB/RIF Assay. In this study, the yielding accuracy of spot and sputum samples were almost similar. This is almost similar to the study done by Tingyu Tang and his colleagues in china in 2017G.C (Tingyu & Fang, 2017).
The study conducted in Uganda in 2017G.C by Amanda J and his colleagues stated that, there was no significance difference in diagnosis result Gene Xpert testing between   salivary and non salivary (good quality) specimens. This was also similar in this research, which shows no relation with quality and consistency for TB detection using Gene Xpert MTB/RIF Assay (Amanda et al., 2017). Another study conducted in Uganda in 2017 by Michael E concluded that spot sample is as good as early morning sputum samples for TB diagnosis. This study also concluded that there is no yielding difference between spot and morning collection methods for diagnosis of TB using Gene Xpert MTB/RIF Assay (Michael et al., 2017). p.4 Among 111 suspected TB patients in this study, 15.3% were positive and 84.7% were negative using morning sputum sample. At same time, using spot sputum samples 13.5% positive and 86.5% were negative. This shows both morning and spot samples have almost similar yielding accuracy using GeneXpert MTB/RIF Assay. It was similar to the study in Kenyan 2016G.C, Gene Xpert testing of spot specimens identified 48 (57.0%) of 84 cases; whereas GeneXpert testing of morning specimens identified 50 (66.0%) of 76 cases were observed (Joseph et al., 2016).

Conclusion
Based on results of this study, yielding accuracy of both morning sputum samples and spot sputum samples is almost similar using Gene Xpert MTB/RIF Assay for detection of MTB. Additionally, the value difference between morning and spot result besides the quality and consistency of samples is not significant.