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Abstract Background When a test for diagnosis of infectious diseases is introduced in a resource-limited setting, monitoring quality is a major concern. An optimized design of experiment and statistical models are required for this assessment. Methods Interferon-gamma release assay to detect tuberculosis (TB) infection from whole blood was tested in Hanoi, Viet Nam. Balanced incomplete block design (BIBD) was planned and fixed-effect models with heterogeneous error variance were used for analysis. In the first trial, the whole blood from 12 donors was incubated with nil, TB-specific antigens or mitogen. In 72 measurements, two laboratory members exchanged their roles in harvesting plasma and testing for interferon-gamma release using enzyme linked immunosorbent assay (ELISA) technique. After intervention including checkup of all steps and standard operation procedures, the second trial was implemented in a similar manner. Results The lack of precision in the first trial was clearly demonstrated. Large within-individual error was significantly affected by both harvester and ELISA operator, indicating that both of the steps had problems. After the intervention, overall within-individual error was significantly reduced (P < 0.0001) and error variance was no longer affected by laboratory personnel in charge, indicating that a marked improvement could be objectively observed. Conclusion BIBD and analysis of fixed-effect models with heterogeneous variance are suitable and useful for objective and individualized assessment of proficiency in a multistep diagnostic test for infectious diseases in a resource-constrained laboratory. The action plan based on our findings would be worth considering when monitoring for internal quality control is difficult on site.

Objectives: In the performance of interferon gamma release assays (IGRA) for the diagnosis of tuberculosis (TB) infection, false-negative results are a major obstacle. In active TB patients, treatment-dependent changes of the negative test results remain unknown., Methods: The treatment course of 19 smear-positive/culture-confirmed TB patients who had IGRA-negative results by QuantiFERON-TB in-tube (QFT-IT) method at the time of diagnosis (month 0) in a previous study, were monitored in the present study. Blood was further collected at months 2 and 7, and the concentrations of 27 immune molecules were measured in the plasma supernatants remaining after performing the IGRA, using a suspension array system., Results: After initiating treatment, eight of the 19 QFT-IT-negative patients showed positive conversion, whereas the remaining 11 (58%) did not; the interferon gamma (IFN-γ) response was restored to levels higher than 1 IU/ml in only three of the eight patients with positive conversion. Plasma concentrations of interleukin 1 receptor antagonist, interleukin 2, and interferon gamma-induced protein 10 remained low after Mycobacterium tuberculosis-specific antigen stimulation at months 2 and 7 in the continuously QFT-IT-negative group, whereas the parameters were elevated only in the transiently QFT-IT-negative group., Conclusions: It was demonstrated that a majority of active TB patients showing negative IGRA results did not regain sufficient levels of immune responsiveness despite successful treatment., (Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objectives: We investigated the relationship between tuberculosis recurrence and Mycobacterium tuberculosis antigen-stimulated interferon-gamma (IFN-γ) responses during treatment., Methods: Plasma IFN-γ levels in active pulmonary tuberculosis patients (n = 407) were analyzed using QuantiFERON-TB Gold In-Tube™ (QFT-IT) at 0, 2, and 7 months of the 8-month treatment received from 2007 to 2009 and the patients were followed up for another 16 months after treatment. Risk factors for recurrence were assessed using the log-rank test and Cox proportional hazard models. Random coefficient models were used to compare longitudinal patterns of IFN-γ levels between groups., Results: QFT-IT showed positive results in 95.6%, 86.2%, and 83.5% at 0, 2, and 7 months, respectively. The antigen-stimulated IFN-γ responses varied significantly during the treatment course (P < 0.0001). Unexpectedly, positive-to-negative conversion of QFT-IT results between 0 and 2 months was significantly associated with earlier recurrence (adjusted hazard ratio, 5.57; 95% confidence interval, 2.28-13.57). Time-dependent changes in IFN-γ levels were significantly different between the recurrence and nonrecurrence groups (P < 0.0001)., Conclusions: Although the IGRA response varies individually, early response during the treatment course may provide an insight into host immune responses underlying tuberculosis recurrence., (Copyright © 2014 The British Infection Association. Published by Elsevier Ltd. All rights reserved.)

Mannose-binding lectin (MBL) binds to pathogens and induces complement-mediated opsonophagocytosis. Although the association between MBL2 polymorphisms and tuberculosis (TB) has been studied in various populations, the results are controversial. We explored the stages of TB associated with MBL2 polymorphisms. X/Y (rs7096206) and A/B (rs1800450) were genotyped in 765 new patients with active pulmonary TB without HIV infection and 556 controls in Hanoi, Viet Nam. The MBL2 nucleotide sequences were further analyzed, and plasma MBL levels were measured in 109 apparently healthy healthcare workers and 65 patients with TB. Latent TB infection (LTBI) was detected by interferon-gamma release assay (IGRA). The YA/YA diplotype, which exhibited high plasma MBL levels, was associated with protection against active TB in younger patients (mean age = 32)≦ 45 years old (odds ratio, 0.61; 95% confidence interval, 0.46-0.80). The resistant diplotype was less frequently found in the younger patients at diagnosis (P = 0.0021). MBL2 diplotype frequencies and plasma MBL levels were not significantly different between the IGRA-positive and -negative groups. MBL2 YA/YA exhibited a protective role against the development of TB in younger patients, whereas the MBL2 genotype and MBL levels were not associated with LTBI. High MBL levels may protect against the early development of pulmonary TB after infection., (Copyright © 2014 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)