ABSTRACT
A comparative study of Immuno-chromatographic test (Serology) with Ziehl-Neelsen (Z-N) staining technique was carried on patients who are suspected of having tuberculosis in the General Hospital, Minna, Niger state, Nigeria. The study was done to compare their performance in the diagnosis of tuberculosis. Three Hundred (300) patients were enlisted for the study but only Two Hundred and Fifty-two (252)patients completed the study. Three (3) Sputum samples were collected from each patient for Z-N staining and 5ml of whole blood was collected from each patient concurrently for serological testing. At the end of the studies the result shows a Raw Agreement of 76% (range of 75-77%), Positive Agreement 1.4% (range of 1.2-1.6%), and a Negative Agreement of 75% (range of 74-76%) between the 2 methods. The Coefficient of Agreement ranged between 0.006 and 0.015(range of κ is 0.006-0.014; range of T is 0.0056- 0.0154). The Sensitivity of the serological test was 12.76%, while it Specificity was 92.42%. It has a Positive Predictive value of 16.22%, while the negative predictive value was 80.53%. With these results the level of Agreement (0.009) between the Immuno-chromatographic Test Strip and Sputum Z-N stain in the diagnosis of PTB is very low and hence Immuno-chromatographic Test Strip cannot replace Z-N staining in the diagnosis of PTB.
CHAPTER ONE
1.0 INTRODUCTION
1.1 Epidemiology
Tuberculosis (TB) is a granulomatous disease caused by Mycobacterium tuberculosis (M. tuberculosis) that affects all organs of the body with varying frequencies (Kumar et al., 2007). The primary mode of infection is by inhalation of droplets laden with the organism (Konstatinos, 2010), hence the vast majority of tuberculosis disease is pulmonary; that is, in the lungs, this accounted for about 75% of all TB infections (WHO, 2007). Other sites of affectations include the Central Nervous System, the Musculo-skeletal System, the Uro-genital system, the Gastrointestinal System and other Systems in the Human body; these other sites outside of the lungs are collectively referred to as extra-pulmonary tuberculosis (Kumar et al., 2007).
A third of the world’s populations are said to be infected with M. tuberculosis, (WHO, 2009). And new infections occur at a rate of about one per second (WHO, 2009). The proportion of people who become sick with tuberculosis each year is stable or falling worldwide but, because of population growth, the absolute number of new cases is still increasing (WHO, 2009).
In 2007 there were an estimated 13.7 million chronic active cases, 9.3 million new cases, and 1.8 million deaths, mostly in developing countries (WHO, 2009). In addition, more people in the developed world are contracting tuberculosis because their immune systems are compromised by immunosuppressive drugs, substance abuse, or Acquired Immune Deficiency Syndrome (AIDS). The distribution of tuberculosis is not uniform across the globe; about 80% of the population in many Asian and African countries test positive in tuberculin tests, while only 5-10% of the US population test positive (Kumar et al., 2007).
About 90% of those infected with Mycobacterium tuberculosis have asymptomatic, latent TB infection (sometimes called LTBI), with only a 10% lifetime chance that a latent infection will progress to TB disease (Kumar et al., 2007). However, if untreated, the death rate for these active TB cases is more than 50% (Onyebujoh, P. & Graham, A. W., 2004).
Accurate identification of LTBI is the key to prevention of the disease among persons at risk (Payam et al., 2006). However, diagnosing this category of Tuberculosis infection i.e. latent Tuberculosis infection, has been a real problem because, hitherto, the modality being used is the Tuberculin Skin Test (TST) using Purified Protein Derivative (PPD) of M. tuberculosis antigen, the response of the host is then measured to determine the state of the patient bearing in mind the risk status (CDC, 2000). However, cross reactivity of this antigen with other strains of the mycobacterium species, especially the Non Tuberculous Mycobacterium (NTM) species raised the issues of false positivity; also previous vaccination with Baccille Calmette-Guerin (BCG) (an attenuated extract of M. bovis) gives a false positive reaction; whilst chronic illnesses including active tuberculosis disease can give a false negative reaction to tuberculin skin test even amongst active sufferers (Pai, 2005).
To resolve these issues, that is, cross reactivity with the Non Tuberculous Mycobacterium (NTM) specie, there is a need to evolve a diagnostic method that will measure response to antigen specific to M. tuberculosis alone, this is what the relatively new Immunologic test for TB infection seeks to resolve. These tests seek to demonstrate the production of Antibody in the infected individual in response to some specific Antigens e.g. Early Secreted Antigen Target 6 kilodalton (ESAT6) and Culture Filtrate Protein 10 kilodalton (CFP10) that are unique to Mycobacteria of the Mycobacterium Tuberculosis Complex (MTC). MTC include M. tuberculosis, M. bovis, M.africanum, M. canetti and M. microti. These antigens are not present in common NTM (e.g. M. avium) as well as BCG vaccination strains (Kanget al., 2007).
ESAT6 and CFP 10 are located on Region of Difference 1 (RD1) on the M. tuberculosis genome hence they are referred to as RD1 based (Payam et al., 2006).
Current evidence on the performance of Interferon gamma (IFN-γ) assays suggests that RD1-based IFN-γ assays have higher specificity, better correlation with surrogate measures of exposure to M. tuberculosis, and less cross-reactivity due to BCG vaccination than the Tuberculin Skin Test (TST) (Payam et al., 2006). RD1-based IFN-γ assays that use a mixture of ESAT-6 and CFP-10 seem to be at least as sensitive as the PPD-based TST in active TB (Payam et al., 2006).
1.2 Statement of the Problem
A third of the World’s population is said to be infected with Mycobacterium tuberculosis (MTB). With about eight Million cases of active disease occurring each year, Tuberculosis (TB) is therefore regarded as a global health problem (WHO, 2007).
Furthermore, the continual migration of people from the developing world with a high incidence of Tuberculosis to highly industrialized countries with less incidence of TB has made MTB a global emergency (Dye et al., 2005).
Also with improvement in the quality of healthcare worldwide, people with chronic illnesses now survive longer albeit, with low immunity, this will enhance the reversion from latent (asymptomatic) MTB infection to active Tuberculosis.
However, despite this prevalence, diagnosis of TB infection in Human has remained a thorny medical issue; all the currently available diagnostic modalities have their drawbacks; Diagnosis of MTB based on Clinical evaluation, Chest radiography and Sputum staining and Microscopy are sensitive but not specific (Anderson et. al., 2000), Culture of bacteria is
time-consuming and the yield is low (Sarmiento et al., 2003), nucleic acid-based methods such as polymerase chain reaction (PCR) are not consistently accurate enough for the diagnosis of smear-negative pulmonary TB (Roba et al., 2010). Tuberculin Skin Test (TST) using Purified Protein Derivative (PPD) largely used for screening has a major drawback of cross-reactivity with antigens derived from several Mycobacteria species, including attenuated M. bovis (Bacille Calmette Guerin (BCG)) used for vaccination, greatly decreasing the specificity of the TST (Roba et al., 2010). Moreover, 10-20% of patient with TB have a negative result with TST due to reduction in their immune competence; this percentage increases with advanced disease or with immunodeficiency due to Human Immune Deficiency Virus (HIV) infections (Converse et al., 1997).
However, it was discovered that MTB infections evoke strong cell-mediated immune response, and detection of specific immune cells might be a mean to detect infection (Delgado et al., 2002).
1.3 Justification for the Research
Immunological tests are Convenient and fast, they can therefore be adopted as a point of care test.
Immunological test do not have the subjectivity inherent in measurements of skin indurations produced by the TST.
Furthermore, there is only one patient’s visit, unlike Ziehl-Neelsen (Z-N) stain/microscopy where three (3) samples collected at three different times are needed per patient.
Also the ability to perform repeat testing without boosting the response, as may occur with serial tuberculin testing, makes serological testing a more attractive option (Payam et al., 2006).
1.4 Aim
To determine if Serological Test (Commercial Immuno-chromatographic Test Strip) can replace Z-N stain and Microscopy in the diagnosis of Tuberculosis.
1.4.1 Specific Objectives
1 Compare the performance of Serological Test using Commercial Immuno- chromatographic Test Strip against Sputum Ziehl-Neelsen (Z-N) stain and Microscopy in the diagnosis of Pulmonary Tuberculosis (PTB) in the Directly Observed Treatment strategy (DOTS) Centre (General Hospital, Minna, Niger State).
1 To find the level of Agreement between the Sputum Z-N stain/Microscopy and Serological Test using Commercial Immuno-chromatographic Test Strip in diagnosis of PTB.
1.5 Study Area
The study was done between February 2011 and August 2011, on 300 patients suspected of having Tuberculosis; they were drawn from the Out-patients Clinics of the General hospital, Minna and patients referred from other Hospitals in Minna and its environs to the Directly Observed Treatment Strategy (DOTS) TB Clinic of the General Hospital, Minna. Minna is the capital city of Niger state, which is one of the States in the North-central zone of Nigeria.
The Hospital is one of the designated centres for the National Leprosy and Tuberculosis programme, where diagnosis of Tuberculosis are done free of charge for all cases. Confirmed cases are also given free treatment courtesy of the Federal Ministry of Health in collaboration with the World Health Organization.
Prior approval of the Ethics and Scientific Committee of the Hospital was sought and permission was granted for the studies.
This material content is developed to serve as a GUIDE for students to conduct academic research
COMPARATIVE STUDY OF COMMERCIAL IMMUNO-CHROMATOGRAPHIC TEST STRIP AGAINST ZIEHL-NEELSEN STAIN IN THE DIAGNOSIS OF TUBERCULOSIS
Project 4Topics Support Team Are Always (24/7) Online To Help You With Your Project
Chat Us on WhatsApp » 09132600555
DO YOU NEED CLARIFICATION? CALL OUR HELP DESK:
09132600555 (Country Code: +234)
YOU CAN REACH OUR SUPPORT TEAM VIA MAIL: [email protected]
09132600555 (Country Code: +234)