Assessment of the Cepheid 3-gene Host Response Fingerstick Blood Test (MTB-HR) on rapid diagnosis of tuberculosis.

ABSTRACTThe World Health Organization has identified high-priority target product profiles for new TB diagnostics which include rapid biomarker-based, non-sputum-based diagnostic testing, using an easily accessible sample. The Cepheid 3-gene Host Response Fingerstick Blood Prototype Test (MTB-HR) quantifies relative mRNA levels of a 3-gene signature (GBP5, DUSP3, and KLF2) from a whole-blood sample on the GeneXpert platform. The objective of the present study was to evaluate the performance of the MTB-HR to distinguish between active tuberculosis (ATB), latent Mycobacterium tuberculosis infection (LTBI), other pulmonary diseases, and healthy volunteers at a tertiary care centre. Among 653 participants enrolled in this study, 192 were diagnosed as having ATB, and the remaining 461 were classified as non-ATB, including 137 cases of LTBI, 224 cases of other pulmonary diseases, and 100 healthy volunteers. The corresponding AUCs of the MTB-HR in distinguishing untreated ATB from non-ATB, LTBI, other pulmonary diseases, and healthy volunteers were 0.814 (95% CI, 0.760-0.868, sensitivity 76.1%, specificity 71.6%), 0.739 (95% CI, 0.667-0.812, sensitivity 59.7%, specificity 78.1%), 0.825 (95% CI, 0.770-0.880, sensitivity 82.1%, specificity 65.6%), 0.892 (95% CI, 0.839-0.945, sensitivity 76.1%, specificity 88.0%), respectively. When only samples with TAT of less than 1 h were included, the AUC of the MTB-HR in distinguishing untreated ATB from non-ATB was largest, 0.920 (95% CI, 0.822-1.000, sensitivity 81.3%, specificity 87.7%). In conclusion, the MTB-HR assay shows potential as a rapid, blood-based screening and triage test for ATB, especially for untreated ATB, with the advantage of increased diagnostic yield since blood is more readily available.

An evolving approach to the laboratory assessment of COVID-19.

As the 2019 novel coronavirus disease (COVID-19) outbreak has evolved in each country, the approach to the laboratory assessment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has had to evolve as well. This review addresses the evolving approach to the laboratory assessment of COVID-19 and discusses how algorithms for testing have been driven, in part, by the demand for testing overwhelming the capacity to accomplish such testing. This review focused on testing in the USA, as this testing is evolving, whereas in China and other countries such as South Korea testing is widely available and includes both molecular testing for SARS-CoV-2 as well as serological testing using both enzyme-linked immunosorbent assay methodology and lateral flow immunoassay methodology. Although commercial testing systems are becoming available, there will likely be insufficient numbers of such tests due to high demand. Serological testing will be the next testing issue as the COVID-19 begins to subside. This will allow immunity testing as well as will allow the parameters of the COVID-19 outbreak to be defined.

Mass spectrometry biotyper system identifies enteric bacterial pathogens directly from colonies grown on selective stool culture media.

We evaluated the performance and cost-effectiveness of a matrix-assisted laser desorption ionization time-of-flight mass spectrometry-based Biotyper system for the routine identification of common enteric bacterial pathogens seen in middle Tennessee from suspicious colonies grown on selective stool culture media. A total of 304 suspicious colonies were selected and further identified from 605 stool specimens. The suspicious colonies were analyzed by the Biotyper system, and the results were compared to those from routine phenotypic methods, which identified 22 Salmonella species, 39 Shigella species, 3 enterohemorrhagic Escherichia coli (EHEC) isolates, 2 Yersinia enterocolitica isolates, 2 Campylobacter species, and 236 gastrointestinal normal flora isolates. The Biotyper system correctly identified the Salmonella species, Yersinia enterocolitica, and Campylobacter species but failed to distinguish the Shigella species and EHEC isolates from E. coli. Among the 236 normal flora isolates, 233 (98.7%) and 228 (96.6%) agreed at the genus and species levels, respectively, between the phenotypic and Biotyper methods. Organism identification scores were insignificantly different between colonies directly from selective media and subsequently from pure subculture. The entire Biotyper identification procedure, from smear preparation to final result reporting, can be completed within 30 min. The Biotyper system provides a rapid and simple screening tool for identifying many, but not all, suspicious colonies grown on selective media within 24 h after inoculation, which shortens test turnaround time by 2 to 3 days.

Assessment of Clostridium difficile infections by quantitative detection of tcdB toxin by use of a real-time cell analysis system.

We explored the use of a real-time cell analysis (RTCA) system for the assessment of Clostridium difficile toxins in human stool specimens by monitoring the dynamic responses of the HS27 cells to tcdB toxins. The C. difficile toxin caused cytotoxic effects on the cells, which resulted in a dose-dependent and time-dependent decrease in cell impedance. The RTCA assay possessed an analytical sensitivity of 0.2 ng/ml for C. difficile toxin B with no cross-reactions with other enterotoxins, nontoxigenic C. difficile, or other Clostridum species. Clinical validation was performed on 300 consecutively collected stool specimens from patients with suspected C. difficile infection (CDI). Each stool specimen was tested in parallel by a real-time PCR assay (PCR), a dual glutamate dehydrogenase and toxin A/B enzyme immunoassay (EIA), and the RTCA assay. In comparison to a reference standard in a combination of the three assays, the RTCA had a specificity of 99.6% and a sensitivity of 87.5% (28 of 32), which was higher than the EIA result (P = 0.005) but lower than the PCR result (P = 0.057). In addition, the RTCA assay allowed for quantification of toxin protein concentration in a given specimen. Among RTCA-positive specimens collected prior to treatment with metronidazole and/or vancomycin, a significant correlation between toxin protein concentrations and clinical CDI severities was observed (R(2) = 0.732, P = 0.0004). Toxin concentrations after treatment (0.89 ng/ml) were significantly lower than those prior to the treatment (15.68 ng/ml, Wilcoxon P = 0.01). The study demonstrates that the RTCA assay provides a functional tool for the potential assessment of C. difficile infections.

Prevalence and management of invalid GeneXpert enterovirus results obtained with cerebrospinal fluid samples: a 2-year study.

A total of 525 cerebrospinal fluid (CSF) samples submitted during the 2007 and 2008 enteroviral seasons were included in a study to determine the prevalence of and potential risk factors for invalid Cepheid GeneXpert enterovirus assay (GXEA) results, as well as possible solutions for the problem. The invalid GXEA results were reported for 43 (8.2%) specimens and correlated with increased visibility of red blood cells (P < 0.0001) but not with CSF xanthochromia and clotting. Invalid GXEA result rates were markedly diminished by 82.1% and 96.0% and test sensitivities were minimally decreased by 1.7% and 3.6% when these specimens were tested at a 1:5 dilution and after a freeze-thaw cycle, respectively.

Comparative evaluation of three commercial systems for nucleic acid extraction from urine specimens.

A nucleic acid extraction system that can handle small numbers of specimens with a short test turnaround time and short hands-on time is desirable for emergent testing. We performed a comparative validation on three systems: the MagNA Pure compact system (Compact), the NucliSens miniMAG extraction instrument (miniMAG), and the BioRobot EZ1 system (EZ1). A total of 75 urine specimens submitted for polyomavirus BK virus detection were used. The human beta-actin gene was detected on 75 (100%), 75 (100%), and 72 (96%) nucleic acid extracts prepared by the miniMAG, EZ1, and Compact, respectively. The miniMAG produced the highest quantity of nucleic acids and the best precision among the three systems. The agreement rate was 100% for BKV detection on nucleic acid extracts prepared by the three extraction systems. When a full panel of specimens was run, the hands-on time and test turnaround time were 105.7 and 121.1 min for miniMAG, 6.1 and 22.6 min for EZ1, and 7.4 and 33.7 min for Compact, respectively. The EZ1 and Compact systems processed automatic nucleic acid extraction properly, providing a good solution to the need for sporadic but emergent specimen detection. The miniMAG yielded the highest quantity of nucleic acids, suggesting that this system would be the best for specimens containing a low number of microorganisms of interest.