Over one million new cases of tuberculosis (TB) and 239,000 TB-related deaths occur in children each year. Young children, especially those with HIV, are more likely to present with disseminated or extrapulmonary TB and paucibacillary disease, often missed by respiratory sampling. Our collaborators have developed methods to rapidly quantify M. tuberculosis (Mtb)-specific antigen (Ag) peptide fragments (CFP-10/ESAT-6) in blood using antibody-labeled and energy-focusing porous discoidal silicon nanoparticles (nanodisks) with high-throughput mass spectrometry (MS) enhancing sensitivity and specificity for TB diagnosis. In pilot studies, NanoDisk-MS was able to detect Mtb-Ag in the blood of HIV-infected adults with sputum culture-confirmed TB, as well as those with culture-negative and extrapulmonary TB. NanoDisk-MS features make it optimal particularly for HIV-infected children including easily collectable small blood volume requirement (<1ml) and potential for detection of paucibacillary and disseminated TB. We propose using archived specimens and data from the Pediatric Urgent Start of HAART (PUSH) Study (NCT02063880) to evaluate the performance of NanoDisk-MS detected Mtb-Ag for diagnosis (Aim 1), mortality prognosis (Aim 2), and treatment response (Aim 3) in HIV-infected children. In addition to assessing conventional diagnostic performance measures, we propose to use advanced epidemiologic methods (Bayesian latent class analysis) given the context of an imperfect reference (Exploratory aim). In the PUSH cohort, 6-month mortality was high (61/100 PY), as was prevalence of confirmed TB (8%) by reference Xpert/culture. Mortality was especially high among the 44% of children who had ?unconfirmed TB? with negative reference Xpert/culture in respiratory samples. We hypothesize NanoDisk-MS will 1) have similar diagnostic performance to the reference Xpert/culture among children with confirmed TB without the need for sputum, 2) identify additional children with TB missed by the reference including those at highest risk of death, and 3) provide a useful surrogate marker of treatment response with significant decline in quantitative levels during successful treatment. While the current iteration of NanoDisk-MS does require resource intensive technology, efforts are underway to develop point-of-care Mtb-Ag detection to increase scalability in resource limited settings. It is important to evaluate pediatric samples throughout this process to confirm diagnostic performance and to guide the move towards point-of-care, to ensure performance is not reduced in children. Using cryopreserved samples from a well-characterized cohort of HIV-infected children who underwent intensive TB evaluation provides an opportunity for efficient evaluation of a novel diagnostic with potential for clinical impact to improve TB diagnosis in HIV-infected children globally.
TB-related morbidity and mortality is high in HIV-infected children who are more likely to present with disseminated or extrapulmonary TB and paucibacilliary disease potentially missed by respiratory sampling. We have developed methods to identify M. tuberculosis (Mtb)-specific antigen (Ag) peptide fragments (CFP- 10/ESAT-6) in easily collected small blood volumes and rapidly quantify their concentrations using antibody- labeled and energy-focusing porous discoidal silicon nanoparticles (nanodisks) with high-throughput mass spectrometry (MS) enhancing sensitivity and specificity (NanoDisk-MS). The proposed study will utilize cryopreserved specimens from a well characterized cohort of hospitalized HIV- infected children who underwent intensive TB investigations to evaluate performance of NanoDisk-MS for TB diagnosis (Aim 1), mortality prognosis (Aim 2), and treatment response (Aim 3), including the use of advanced epidemiologic methods (Bayesian latent class analysis) in the setting of an imperfect reference (Exploratory Aim).
