Xpert Ultra testing of blood in severe HIV-associated tuberculosis to detect and measure Mycobacterium tuberculosis blood stream infection: a diagnostic and disease biomarker cohort study

Serial Measurement of <i>M. Tuberculosis</i> in Blood from Critically-Ill Patients with HIV-Associated Tuberculosis

“…28 In support of a causal relationship, we found that blood bacillary load measured by the novel methods described had a dose-response relationship with clinical phenotype and mortality, adding to previous evidence. [8] Two attempts at quantifying MTBBSI have been previously reported, both using lysis-centrifugation colonyforming unit (CFU) counting on pre-treatment blood samples. 29,30 31 This study reports the first successful use of microscopy on blood, with bacilli identified in 68 out of 99 total patient samples examined.…”

“…12 Secondly, a lysis-wash method for removal of PCR inhibitors was developed, allowing blood to be tested on the GeneXpert Ò MTB/RIF Ultra platform (Xpert-ultra, Cepheid, Sunnyvale, California) with high sensitivity. 8 Third, to allow culture of blood after initiation of ATT, we reduced antimicrobial carry-over by removing plasma or performing lysis-wash preprocessing of blood.…”

“…Batched samples were thawed and tested using Xpert-Ultra cartridges. 8 Full results were output from the Xpert software as .txt files and cycle threshold (Ct) values extracted using a custom R script. The mean Ct value of the four rpoB probes was used as the summary value; for rpoB probe negative / IS1081-IS6110-positive samples ("trace" positive), we used the IS1081-IS6110 Ct values to impute mean rpoB probe Ct value.…”

“…The mean Ct value of the four rpoB probes was used as the summary value; for rpoB probe negative / IS1081-IS6110-positive samples ("trace" positive), we used the IS1081-IS6110 Ct values to impute mean rpoB probe Ct value. 8 Myco/F lytic blood cultures Myco/F Lytic cultures inoculated as above were transferred to the National Health Laboratory Service (NHLS) and incubated within 24 h of venesection in the automated BACTEC system. Time-to-positivity (TTP) was extracted from BACTEC software.…”

“…5 A dose-response relationship exists between blood bacillary load, clinical and immunological phenotype, and risk of death. 7,8 Apart from diagnostic trials, 9 severe HIV-associated TB has not been the focus of dedicated interventional trials of therapies to reduce mortality. Adequacy of anti-TB treatment (ATT) in HIV-associated disease has been tested predominantly in smear-positive, ambulant outpatients with pulmonary TB, and not in critically-ill inpatients with MTBBSI.…”