Improving TB Diagnosis and Treatment Through Basic, Applied and Health Systems Research
Improving TB Diagnosis and Treatment Through Basic, Applied and Health Systems Research
TB is a global health problem and in South Africa rates as the second most important problem
 in terms of Burden of Disease. There are many reasons for this, among which are diagnostic
 difficulties, extended treatments, drug resistance and health care provision. This
 application is concerned with all these drivers and will focus activities on a clinic which
 provides basic care in a very deprived socio-economic area of greater Cape Town, South
 Africa.

 Patients studied in routine, but demanding environments are our focus as these clinics are
 representative of many areas where TB (and HIV) are found at high prevalence. If the
 constraints of working in such areas can be understood and appropriate changes that work
 made, the investigators believe the outputs and policy changes generated in this study will
 contribute to future success in other settings.

 The investigators wish to study the implementation of the Xpert®MTB/RIF (Xpert) and Xpert
 Ultra (Ultra) systems in situ using a randomised controlled trial design, as opposed to a
 remote site (central laboratory), to assess whether time to diagnosis can be improved using
 point of care (POC). The investigators wish to maximise this opportunity by collecting
 biological samples from a patient population experiencing a TB epidemic for the evaluation of
 future TB diagnostics.

 Using human DNA, the investigators will attempt to determine reasons for poor or no treatment
 response. Two possibilities exist for this: a) the M. tuberculosis strain is resistant to the
 drug in question or b) the patient is highly susceptible to the bacterium. The investigators
 will determine the exome sequences of study participants with susceptible M. tuberculosis
 strains who show poor or no response, and compare this with rapid responders. Using 16S rRNA
 sequencing, the investigators will also observe how the microbiome of TB patients is altered
 during TB treatment and how this is associated with treatment outcome, as well as after TB
 treatment.

 This project will set the foundation for the implementation of new POC TB diagnostic
 technologies in clinics in South Africa. The biobanked specimens collected can be rapidly
 utilised for nascent technologies. Studying the patient microbiome will provide insights into
 what makes some patients more susceptible to TB and what microbiological changes occur during
 the course of anti-TB treatment.
In 2015, the World Health Organization (WHO) estimated that there were there were an
 estimated 10.4 million new TB cases, 1.8 million deaths, and 580 000 cases of drug-resistant
 TB with 1.4 million deaths from TB among human immunodeficiency virus (HIV) negative people,
 and an additional 0.40 million deaths from HIV-associated TB. The African Region has
 approximately one quarter of the world's cases, and the highest rates of cases and deaths
 relative to population. The proportion of TB cases co-infected with HIV was highest in
 countries in the African Region; overall, 33% of TB cases were estimated to be co-infected
 with HIV in this region. South Africa is one of the countries with the highest number of
 incident cases in 2011 (410,000 - 600,000) making it one of the 22 high burden countries.
 Current estimates suggest that 330,000 of the incident cases are co-infected with HIV.
 Despite being declared an emergency in 1996 and the implementation of Direct Observed Therapy
 Short courses (DOTS) the case detection rate remains below the WHO target of 85%.

 The DOTS strategy is promoted by the WHO as the optimal program to control the TB epidemic.
 According to the WHO, antibiotic treatment is effective in about 90% of patient, however, the
 duration of treatment places an enormous burden on health care services and patients often
 stop taking the treatment once the symptoms have improved. This implies that it is extremely
 challenging for TB control programmes to reach the WHO's target of 85% successful treatment.
 It is hypothesized that shortening the duration of treatment will significantly impact on
 adherence as well as treatment outcome. However therapeutic options are extremely limited
 given that there has been little antibiotic development over the past 40 years. The situation
 with respect to antibiotic resistant TB is considerably worse. Recent molecular
 epidemiological studies have provided convincing evidence that MDR-TB in South Africa is
 caused mostly by the transmission of MDR strains, as demonstrated by well-documented clonal
 outbreaks and elevated rates of primary resistance (in some places as high as 80%) among
 MDR-TB cases.

 One of the effective means of combating drug resistant TB is the rapid diagnosis of any
 person with TB and detection of any potentially drug resistant TB strains in the early stages
 of the disease. Smear microscopy remains the state of the art diagnostic in many developing
 countries, but this method shows poor sensitivity which is further compromised by HIV
 co-infection. Culture-based methods remain the gold standard but are time consuming often
 leading to extensive diagnostic delays.

 The WHO has called for expanding access to M. tuberculosis culture and drug susceptibility
 testing (DST) as part of the WHO Global laboratory strategic plan. Accordingly, 50% of the
 global population should have gained access to culture and DST services by 2010 and the
 scale-up should be completed by 2015, covering more than 5 billion people. But even if the
 access to culture and culture-based DST can be expanded in resource-poor settings, the slow
 turn-around time (several weeks) of culture-based DST limits the impact it has on treatment
 decisions and patient outcomes. This is especially true for HIV-infected TB suspects and
 children in whom a treatment decision should not be delayed due to rapid progression of
 disease and high risk of early mortality.

 In June 2008, the WHO recommended the use of line probe assay (LPA) for rapid detection of
 MDR-TB in smear positive sputum specimens. These LPAs use polymerase chain reaction (PCR) to
 amplify regions of interest in the mycobacterial genome followed by reverse hybridization to
 sequence specific probes. The most prominent commercial DST product is the GenoType®
 MTBDRplus assay (HAIN Lifescience, GmbH, Nehren, Germany) for use in smear positive sputum
 samples or culture isolates. Both methods detect M. tuberculosis complex infection as well as
 mutations which cause RIF resistance (rpoB gene). The MTBDRplus assay can detect INH
 resistance by analysing the katG gene and the inhA promoter gene region. The sensitivity and
 specificity to detect INH and RIF resistance using the Genotype MTBDRplus has been shown to
 be high. In 2013, the Genotype MTBDRsl was endorsed by the WHO. This second line LPA assay
 allows the detection of resistance to fluoroquinolones, aminoglycosides, CAP, and EMB by
 targeting the gyrA, rrs and embB genes in M. tuberculosis, respectively. The sensitivity of
 the Genotype MTBDRsl ranges from 75 to 91% for detection of fluoroquinolone resistance, 77 to
 85% for KANA resistance, 80 to 87% for CAP resistance, and 57 to 69% for EMB resistance.

 There are some limitations to this technology. Firstly, not all the genotypes conferring
 resistance are known and secondly, DST methods are not all standardized and vary in their
 performance. This may result in suboptimal and variable sensitivities of the molecular based
 methods for certain drugs. The open-tube format of the method also allows for potential
 amplicon cross-contamination, which may lead to incorrect diagnoses and subsequent
 mismanagement of patients. Thus expensive infrastructure is required to enable these tests to
 be run.

 An alternative diagnostics and DST solution became available in December 2010, when the WHO
 recommended the use of Xpert®MTB/RIF (Xpert), a closed, fully integrated and automated nested
 real-time PCR assay for rapid and simultaneous detection of M. tuberculosis complex and RIF
 resistance, for use as the first diagnostic test in TB suspects at risk of MDR-TB and
 HIV-associated TB. The method allows sample decontamination, PCR and real-time analysis to
 occur within a single disposable plastic cartridge. The turnaround time is less than 2 hours.
 The Xpert performs well for smear positive specimens, and shows promise for improving the
 diagnosis of smear-negative and extrapulmonary TB and RIF resistant TB specimens. Thus, this
 system may prove especially useful in settings where co-infection rates with HIV are high.
 However, the Xpert assay only has a ~70% sensitivity for detecting TB in smear negative
 samples and only provides information on RIF resistance. Furthermore, if the test indicates
 RIF resistance the specimen needs to be sent for second-line DST. Thus, the time gained by
 the rapid test is lost by the need for conventional DST tests. In addition, the test does not
 provide pharmacogenetic information which may be essential for formulation of an appropriate
 treatment regimen. Recently, the WHO has endorsed Xpert Ultra (Ultra), the successor of
 Xpert®MTB/RIF. Ultra retains its ability to detect rifampicin resisitance and is reported to
 be ten times more sensitive than its Xpert®MTB/RIF predecessor with total run-time for the
 nearly halved.

 Xpert and MDRplus have been successfully implemented in the South African National Health
 Laboratory Service (NHLS). The NHLS implemented MTBDRplus LPA in 2008 with the aim to reduce
 the time to the initiation of treatment as compared to culture-based diagnostics. Despite a
 laboratory turnaround time of <7 days, the time to initiation of treatment was on average 55
 days when MDR-TB was diagnosed. Examination of the cause of the additional delay showed that
 the laboratory turnaround time was significantly reduced but remained at 22 days for smear
 positive samples and 29 days for smear negative samples. This was explained by the high
 volume of samples needing testing and subsequent evaluation of the tests before distribution
 to the clinics. This highlights the need for re-tooling of the diagnostic laboratory to
 rapidly process and disseminate genetic-based tests. In addition, this study also highlighted
 the need to improve health systems to respond to rapid genetic based tests. No change was
 observed in the time between sending the diagnostic report and initiation of treatment
 (average 21 days, irrespective of the diagnostic method.

 It is known that diagnostics in TB is difficult, but central to the control programme.
 Despite recent advances, diagnostics are not yet sufficiently rapid, cheap and comprehensive
 and even the introduction of the Xpert instrument has not dramatically reduced time to
 diagnosis or presumably patient retention. In South Africa, Xpert was introduced and scaled
 up rapidly in an attempt to address the TB epidemic, however, the testing equipment is
 centrally located.

 The investigators propose a study which implements Xpert®MTB/RIF and Xpert Ultra for the
 testing of TB in symptomatic patients at the POC which could potentially improve the
 time-to-diagnosis and thus improve time to treatment initiation compared to centralised
 testing.
Inclusion Criteria:

 -  The patient is willing to provide specimens (urine, blood, stool, respiratory tract
 specimens and swabs)

 -  The patient is clinically suspected of having TB (Two WHO TB symptoms for HIV negative
 patients and one WHO TB symptom if HIV positive)

 Exclusion Criteria:

 -  The patient is under the age of 18 years old

 -  The patient declines consent

 -  The patient has too few clinical symptoms for TB