Detection of Dengue Virus in Mosquito Extracts and Human Clinical Samples Using a Field Expedient Molecular Platform.

Dengue fever occurs in localized outbreaks and can significantly erode troop strength and mission readiness. Timely identification of dengue virus (DENV) provides for rapid and appropriate patient management decisions, such as medical evacuation and supportive therapies, as well as help to promote Force Health Protection through vector control and personal protective measures. The "Ruggedized" Advanced Pathogen Identification Device is a field-friendly PCR (Polymerase Chain Reaction) platform that can be used to facilitate early identification of DENV. We developed a dry-format PCR assay on this platform. The assay demonstrated 100% analytical specificity for detecting dengue using a cross-reactivity panel. We used a panel of 102 acute, DENV isolation positive serum samples and 25 DENV negative samples; the assay demonstrated a clinical sensitivity of 97.1% (95% C.I. 91.6-99.4%) and specificity of 96.0% (95% C.I. 79.7-99.9%) in identifying patients with dengue infection. We also used the assay to test mosquito homogenates from 28 adult female Aedes aegypti. A single DENV infected mosquito was identified using the PCR assay and confirmed using immunofluorescence as a reference method. Much of the testing was performed under austere field conditions. Together, our results demonstrate the utility of this assay for detecting DENV in vector and human samples in field environments.

Within-subject interlaboratory variability of QuantiFERON-TB gold in-tube tests.

BACKGROUND: The QuantiFERON®-TB Gold In-Tube test (QFT-GIT) is a viable alternative to the tuberculin skin test (TST) for detecting Mycobacterium tuberculosis infection. However, within-subject variability may limit test utility. To assess variability, we compared results from the same subjects when QFT-GIT enzyme-linked immunosorbent assays (ELISAs) were performed in different laboratories. METHODS: Subjects were recruited at two sites and blood was tested in three labs. Two labs used the same type of automated ELISA workstation, 8-point calibration curves, and electronic data transfer. The third lab used a different automated ELISA workstation, 4-point calibration curves, and manual data entry. Variability was assessed by interpretation agreement and comparison of interferon-γ (IFN-γ) measurements. Data for subjects with discordant interpretations or discrepancies in TB Response >0.05 IU/mL were verified or corrected, and variability was reassessed using a reconciled dataset. RESULTS: Ninety-seven subjects had results from three labs. Eleven (11.3%) had discordant interpretations and 72 (74.2%) had discrepancies >0.05 IU/mL using unreconciled results. After correction of manual data entry errors for 9 subjects, and exclusion of 6 subjects due to methodological errors, 7 (7.7%) subjects were discordant. Of these, 6 (85.7%) had all TB Responses within 0.25 IU/mL of the manufacturer's recommended cutoff. Non-uniform error of measurement was observed, with greater variation in higher IFN-γ measurements. Within-subject standard deviation for TB Response was as high as 0.16 IU/mL, and limits of agreement ranged from -0.46 to 0.43 IU/mL for subjects with mean TB Response within 0.25 IU/mL of the cutoff. CONCLUSION: Greater interlaboratory variability was associated with manual data entry and higher IFN-γ measurements. Manual data entry should be avoided. Because variability in measuring TB Response may affect interpretation, especially near the cutoff, consideration should be given to developing a range of values near the cutoff to be interpreted as "borderline," rather than negative or positive.