scholarly journals Clinical Evaluation of a Blood Assay to Diagnose Paucibacillary Tuberculosis via Bacterial Antigens

2018 ◽  
Vol 64 (5) ◽  
pp. 791-800 ◽  
Author(s):  
Chang Liu ◽  
Christopher J Lyon ◽  
Yang Bu ◽  
Zaian Deng ◽  
Elisabetta Walters ◽  
...  
Keyword(s):  
Direct Detection ◽  
Clinical Performance ◽  
Chinese Adults ◽  
Mycobacterial Culture ◽  
Clinical Sensitivity ◽  
Disease Site ◽  
Early Secretory Antigenic Target ◽  
Tb Diagnostics ◽  
Culture Positive ◽  
Culture Negative

Abstract BACKGROUND The diagnosis of active tuberculosis (TB) cases primarily relies on methods that detect Mycobacterium tuberculosis (Mtb) bacilli or their DNA in patient samples (e.g., mycobacterial culture and Xpert MTB/RIF assays), but these tests have low clinical sensitivity for patients with paucibacillary TB disease. Our goal was to evaluate the clinical performance of a newly developed assay that can rapidly diagnose active TB cases by direct detection of Mtb-derived antigens in patients' blood samples. METHODS Nanoparticle (NanoDisk)-enriched peptides derived from the Mtb virulence factors CFP-10 (10-kDa culture factor protein) and ESAT-6 (6-kDa early secretory antigenic target) were analyzed by high-throughput mass spectrometry (MS). Serum from 294 prospectively enrolled Chinese adults were analyzed with this NanoDisk-MS method to evaluate the performance of direct serum Mtb antigen measurement as a means for rapid diagnosis of active TB cases. RESULTS NanoDisk-MS diagnosed 174 (88.3%) of the study's TB cases, with 95.8% clinical specificity, and with 91.6% and 85.3% clinical sensitivity for culture-positive and culture-negative TB cases, respectively. NanoDisk-MS also exhibited 88% clinical sensitivity for pulmonary and 90% for extrapulmonary TB, exceeding the diagnostic performance of mycobacterial culture for these cases. CONCLUSIONS Direct detection and quantification of serum Mtb antigens by NanoDisk-MS can rapidly and accurately diagnose active TB in adults, independent of disease site or culture status, and outperform Mycobacterium-based TB diagnostics.

scholarly journals Direct Detection of Shigella in Stool Specimens by Use of a Metagenomic Approach

2017 ◽  
Vol 56 (2) ◽  
Author(s):  
Jie Liu ◽  
Mathieu Almeida ◽  
Furqan Kabir ◽  
Sadia Shakoor ◽  
Shahida Qureshi ◽  
...  
Keyword(s):  
Direct Detection ◽  
Positive Culture ◽  
Accurate Method ◽  
Diagnostic Methods ◽  
Metagenomic Sequencing ◽  
Illumina Hiseq ◽  
Content Type ◽  
Sequence Composition ◽  
Culture Positive ◽  
Culture Negative

ABSTRACTThe underestimation ofShigellaspecies as a cause of childhood diarrhea disease has become increasingly apparent with quantitative PCR (qPCR)-based diagnostic methods versus culture. We sought to confirm qPCR-based detection ofShigellavia a metagenomics approach. Three groups of samples were selected from diarrheal cases from the Global Enteric Multicenter Study: nineShigellaculture-positive and qPCR-positive (culture+qPCR+) samples, nine culture-negative but qPCR-positive (culture−qPCR+) samples, and nine culture-negative and qPCR-negative (culture−qPCR−) samples. Fecal DNA was sequenced using paired-end Illumina HiSeq, whereby 3.26 × 108± 5.6 × 107high-quality reads were generated for each sample. We used Kraken software to compare the read counts specific to “Shigella” among the three groups. The proportions ofShigella-specific nonhuman sequence reads between culture+qPCR+(0.65 ± 0.42%) and culture−qPCR+(0.55 ± 0.31%) samples were similar (Mann-Whitney U test,P= 0.627) and distinct from the culture−qPCR−group (0.17 ± 0.15%,P< 0.05). The read counts of sequences previously targeted byShigella/enteroinvasiveEscherichia coli(EIEC) qPCR assays, namely,ipaH,virA,virG,ial,ShET2, andipaH3, were also similar between the culture+qPCR+and culture−qPCR+groups and distinct from the culture−qPCR−groups (P< 0.001). Kraken performed well versus other methods: its precision and recall ofShigellawere excellent at the genus level but variable at the species level. In summary, metagenomic sequencing indicates thatShigella/EIEC qPCR-positive samples are similar to those ofShigellaculture-positive samples inShigellasequence composition, thus supporting qPCR as an accurate method for detectingShigella.

scholarly journals Evaluation of the Xpert MTB/RIF Ultra Assay for Direct Detection ofMycobacterium tuberculosisComplex in Smear-Negative Extrapulmonary Samples

2018 ◽  
Vol 56 (9) ◽  
Author(s):  
Daniel Perez-Risco ◽  
David Rodriguez-Temporal ◽  
Ivan Valledor-Sanchez ◽  
Fernando Alcaide
Keyword(s):  
Nontuberculous Mycobacteria ◽  
Direct Detection ◽  
Extrapulmonary Tuberculosis ◽  
High Sensitivity ◽  
Rapid Diagnosis ◽  
Clinical Samples ◽  
Content Type ◽  
Smear Negative ◽  
Culture Positive ◽  
Culture Negative

ABSTRACTThe rapid detection ofMycobacterium tuberculosiscomplex (MTUBC) in clinical samples is essential for successful treatment. New techniques such as real-time PCR have been developed in order to facilitate rapid diagnosis, but their sensitivity is low in extrapulmonary specimens, due to the low bacillary load in such samples. A next-generation assay has recently been developed to try to overcome this limitation. The aim of this study was to analyze the effectiveness of the Xpert MTB/RIF Ultra (GX-Ultra) for the detection of MTUBC DNA in 108 smear-negative extrapulmonary specimens that were MTUBC culture positive. In addition, 40 extrapulmonary culture-negative samples and 20 samples with nontuberculous mycobacteria were tested to evaluate the specificity of the assay. All samples were collected between May 1999 and May 2017. The GX-Ultra detected DNA of MTUBC in 82 extrapulmonary specimens that were MTUBC culture positive (75.9% sensitivity; 95% confidence interval [CI], 66.6 to 83.4%). The assay was negative for all clinical specimens that were MTUBC culture negative and the samples with nontuberculous mycobacteria (100% specificity). Furthermore, two (1.8%) samples presented mutations related to rifampin resistance. The highest sensitivity was obtained in samples of lymph nodes (94.1%) and nonsterile fluids (93.7%), followed by tissue specimens (86.6%), stool material (80%), abscess aspirates (64.7%), and sterile fluids (60.5%). Pleural fluids, one of the least optimal samples for detecting DNA of MTUBC, were GX-Ultra positive in 10/21 (47.6%) of cases. In summary, GX-Ultra showed excellent specificity and high sensitivity in paubacillary specimens, making it a useful tool for rapid diagnosis of extrapulmonary tuberculosis.

scholarly journals Direct Detection and Identification of Prosthetic Joint Pathogens in Synovial Fluid (SF) by Metagenomic Shotgun Sequencing

2017 ◽  
Vol 4 (suppl_1) ◽  
pp. S32-S32
Author(s):  
Morgan Ivy ◽  
Matthew Thoendel ◽  
Patricio Jeraldo ◽  
Kerryl Greenwood-Quaintance ◽  
Arlen D Hanssen ◽  
...  
Keyword(s):  
Synovial Fluid ◽  
Infectious Diseases ◽  
Direct Detection ◽  
Marker Gene ◽  
Shotgun Sequencing ◽  
Detection And Identification ◽  
Review Course ◽  
Board Review ◽  
Culture Positive ◽  
Culture Negative

Abstract Background Detection and identification of microorganism(s) involved in periprosthetic joint infection (PJI) can inform surgical management and directed antibiotic therapy. Metagenomic shotgun sequencing is a powerful tool with the potential to change how many PJIs are diagnosed as it allows direct detection and identification of pathogens in clinical specimens. In the largest series to date, we utilized a metagenomics-based approach applied to SF to define potential microbial etiologies of failed total knee arthroplasties (TKAs). Methods Synovial fluid was collected from 112 failed TKAs [74 PJI and 38 aseptic implant failure (AF)] via preoperative arthrocentesis. Cell count and differential, standardized culture and DNA-based metagenomic shotgun sequencing were performed. Human DNA was depleted using the MolYsis basic kit prior to DNA extraction, whole genome amplification, and sequencing. Taxonomic assignment of reads and pathogen identification was achieved using a pipeline incorporating k-mer- and marker gene-based classification software. A scheme for analysis and filtration of false-positives was created and applied, incorporating cut-offs for the number of reads, quality scores, and coverage across a reference genome. Patients were classified as having PJI using the IDSA criteria and expert review. Analyses were recorded as percent agreement, with 95% confidence intervals (CI), of metagenomics to SF culture. Results Metagenomic analysis identified the known pathogen in 54 (90%) (CI, 79.5%–96.2%) of the 60 culture-positive PJIs analyzed and one (2%) (CI, 0.0%–8.9%) potential polymicrobial infection not detected by culture. For the 14 culture-negative PJIs tested, metagenomics showed 79% (CI, 49.2%–95.3%) agreement for negative findings; potential pathogens were identified in three (21%) (CI, 4.7%–50.8%) culture-negative PJI cases, with one being polymicrobial. Of the 37 culture-negative AF cases, metagenomics showed 97% (CI, 85.8%–99.9%) agreement with negative culture and identified one (3%) (CI, 0.0%–14.2%) potential pathogen. For the one culture-positive AF case, metagenomic results were negative, suggesting possible culture contamination. Conclusion Metagenomic shotgun sequencing performed on SF can be used to diagnose PJI and may be particularly useful for culture-negative PJI. Disclosures R. Patel, ASM: Board Member, None; CD Diagnostics, BioFire, Curetis, Merck, Hutchison Biofilm Medical Solutions, Accelerate Diagnostics, Allergan, and The Medicines Company: Grant Investigator, Grant recipient; Curetis: Consultant, Monies paid to my employer; A patent on Bordetella pertussis/parapertussis PCR issued, a patent on a device/method for sonication with royalties paid by Samsung to Mayo Clinic, and a patent on an anti-biofilm substance issued: Patents, Patents, any money is paid to my employer; Actelion: DSMB, Money paid to my employer; ASM and IDSA: Editor’s stipends, Editor’s stipends; NBME, Up-to-Date and the Infectious Diseases Board Review Course: NBME, Up-to-Date and the Infectious Diseases Board Review Course, Honoraria; Roche, ASM, and IDSA: Travel reimbursement, Travel reimbursement

scholarly journals Retrospective diagnosis of lymphatic tuberculosis in frozen samples using two genetic amplification methods, Xpert® MTB/RIF ULTRA and Abbott RealTime MTB Assay

2021 ◽  
Author(s):  
Mariana Fernandez-Pittol ◽  
◽  
Yuliya Zboromyrska ◽  
Angely Román ◽  
Griselda Tudó Vilanova ◽  
...  
Keyword(s):  
Gene Amplification ◽  
Sensitivity And Specificity ◽  
Retrospective Diagnosis ◽  
Mycobacterial Culture ◽  
Initial Culture ◽  
Secondary Objective ◽  
Mycobacterium Avium Intracellulare ◽  
Genetic Amplification ◽  
Culture Positive ◽  
Culture Negative

Objectives. The main objective of the present study is to assess the sensitivity and specificity of a retrospective diagnostic of lymphatic tuberculosis (LTB), testing frozen samples using gene amplification PCR methods. The secondary objective was to compare the results of two different commercial tuberculosis gene amplification methods for this purpose. Material and methods. We retrospectively studied 38 frozen samples, previously processed for mycobacterial culture between January 2014 and August 2019. The results of the previous cultures were: 21 samples positive for Mycobacterium tuberculosis complex (MTB) (5 being smear positive), 7 samples culture positive for Mycobacterium avium-intracellulare complex and 10 samples which were mycobacterial culture negative and discarded for LTB diagnosis, used as controls. The samples were processed using two gene amplification methods: Xpert® MTB/RIF Ultra (Cepheid) and Abbott RealTime MTB Assay (Abbott). Results. Compared to initial culture results the sensitivity and specificity of Xpert® MTB/RIF Ultra were 57.1% and 100% and 52.3 % and 92.5%, respectively for the Abbott RealTime MTB assay. The differences were not statiscally significant. In addition, there were no differences according to the period of freezing. Conclusions. Gene amplification of frozen samples confirmed the diagnosis of lymphatic TB in almost 60% of cases, allowing retrospective diagnosis in initially non suspected cases. Both gene amplification techniques tested were equally useful.

scholarly journals Bacteremic sepsis leads to higher mortality when adjusting for confounders with propensity score matching

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Lisa Mellhammar ◽  
Fredrik Kahn ◽  
Caroline Whitlow ◽  
Thomas Kander ◽  
Bertil Christensson ◽  
...  
Keyword(s):  
Intensive Care ◽  
Clinical Review ◽  
Pathogen Detection ◽  
Mortality Rates ◽  
Blood Cultures ◽  
Population Heterogeneity ◽  
Sepsis Diagnosis ◽  
Culture Positive ◽  
Culture Negative ◽  
Microbial Samples

AbstractOne can falsely assume that it is well known that bacteremia is associated with higher mortality in sepsis. Only a handful of studies specifically focus on the comparison of culture-negative and culture-positive sepsis with different conclusions depending on study design. The aim of this study was to describe outcome for critically ill patients with either culture-positive or -negative sepsis in a clinical review. We also aimed to identify subphenotypes of sepsis with culture status included as candidate clinical variables. Out of 784 patients treated in intensive care with a sepsis diagnosis, blood cultures were missing in 140 excluded patients and 95 excluded patients did not fulfill a sepsis diagnosis. Of 549 included patients, 295 (54%) had bacteremia, 90 (16%) were non-bacteremic but with relevant pathogens detected and in 164 (30%) no relevant pathogen was detected. After adjusting for confounders, 90-day mortality was higher in bacteremic patients, 47%, than in non-bacteremic patients, 36%, p = 0.04. We identified 8 subphenotypes, with different mortality rates, where pathogen detection in microbial samples were important for subphenotype distinction and outcome. In conclusion, bacteremic patients had higher mortality than their non-bacteremic counter-parts and bacteremia is more common in sepsis when studied in a clinical review. For reducing population heterogeneity and improve the outcome of trials and treatment for sepsis, distinction of subphenotypes might be useful and pathogen detection an important factor.

scholarly journals Culture-negative versus culture-positive in pyogenic spondylitis and analysis of risk factors for relapse

2021 ◽  
pp. 1-5
Author(s):  
Guohua Dai ◽  
Shuzhong Li ◽  
Chuqiang Yin ◽  
Yuanliang Sun ◽  
Jianwen Hou ◽  
...  
Keyword(s):  
Risk Factors ◽  
Pyogenic Spondylitis ◽  
Culture Positive ◽  
Culture Negative

scholarly journals Association Between Diabetic Foot Infection Wound Culture Positivity and 1-Year Admission for Invasive Infection: A Multi-Center Cohort Study

2021 ◽  
Author(s):  
Westyn Branch-Elliman ◽  
Daniel Sturgeon ◽  
Adolf W Karchmer ◽  
Hillary J Mull
Keyword(s):  
Diabetic Foot ◽  
Systemic Infection ◽  
Causative Organism ◽  
Invasive Infection ◽  
Diabetic Foot Ulcers ◽  
Diabetic Foot Infections ◽  
Culture Positivity ◽  
Wound Culture ◽  
Culture Positive ◽  
Culture Negative

Abstract Inpatients with culture-positive diabetic foot infections are at elevated risk for subsequent invasive infection with the same causative organism. In outpatients with index diabetic foot ulcers, we found that wound culture positivity was independently associated with increased odds of 1-year admission for systemic infection when compared to culture-negative wounds.

scholarly journals Evaluation of the performance of clinical predictors in estimating the probability of pulmonary tuberculosis among smear-negative cases in Northern Ethiopia: a cross-sectional study

BMJ Open ◽  
2020 ◽  
Vol 10 (11) ◽  
pp. e037913
Author(s):  
Mala George ◽  
Geert-Jan Dinant ◽  
Efrem Kentiba ◽  
Teklu Teshome ◽  
Abinet Teshome ◽  
...  
Keyword(s):  
Pulmonary Tuberculosis ◽  
Cross Sectional Study ◽  
Clinical Suspicion ◽  
Secondary Outcome ◽  
Northern Ethiopia ◽  
Sectional Study ◽  
Cross Sectional ◽  
Smear Negative ◽  
Culture Positive ◽  
Culture Negative

ObjectivesTo evaluate the performance of the predictors in estimating the probability of pulmonary tuberculosis (PTB) when all versus only significant variables are combined into a decision model (1) among all clinical suspects and (2) among smear-negative cases based on the results of culture tests.DesignA cross-sectional study.SettingTwo public referral hospitals in Tigray, Ethiopia.ParticipantsA total of 426 consecutive adult patients admitted to the hospitals with clinical suspicion of PTB were screened by sputum smear microscopy and chest radiograph (chest X-ray (CXR)) in accordance with the Ethiopian guidelines of the National Tuberculosis and Leprosy Program. Discontinuation of antituberculosis therapy in the past 3 months, unproductive cough, HIV positivity and unwillingness to give written informed consent were the basis of exclusion from the study.Primary and secondary outcome measuresA total of 354 patients were included in the final analysis, while 72 patients were excluded because culture tests were not done.ResultsThe strongest predictive variables of culture-positive PTB among patients with clinical suspicion were a positive smear test (OR 172; 95% CI 23.23 to 1273.54) and having CXR lesions compatible with PTB (OR 10.401; 95% CI 5.862 to 18.454). The regression model had a good predictive performance for identifying culture-positive PTB among patients with clinical suspicion (area under the curve (AUC) 0.84), but it was rather poor in patients with a negative smear result (AUC 0.64). Combining all the predictors in the model compared with only the independent significant variables did not really improve its performance to identify culture-positive (AUC 0.84–0.87) and culture-negative (AUC 0.64–0.69) PTB.ConclusionsOur finding suggests that predictive models based on clinical variables will not be useful to discriminate patients with culture-negative PTB from patients with culture-positive PTB among patients with smear-negative cases.

Circulating microbial cell-free DNA is associated with inflammatory host-responses in severe pneumonia

Thorax ◽  
2021 ◽  
pp. thoraxjnl-2020-216013
Author(s):  
Haopu Yang ◽  
Ghady Haidar ◽  
Nameer S Al-Yousif ◽  
Haris Zia ◽  
Daniel Kotok ◽  
...  
Keyword(s):  
Large Scale ◽  
Inflammatory Responses ◽  
Severe Pneumonia ◽  
Systemic Circulation ◽  
Microbial Cell ◽  
Metagenomic Sequencing ◽  
Cell Free Dna ◽  
Free Dna ◽  
Culture Positive ◽  
Culture Negative

Host inflammatory responses predict worse outcome in severe pneumonia, yet little is known about what drives dysregulated inflammation. We performed metagenomic sequencing of microbial cell-free DNA (mcfDNA) in 83 mechanically ventilated patients (26 culture-positive, 41 culture-negative pneumonia, 16 uninfected controls). Culture-positive patients had higher levels of mcfDNA than those with culture-negative pneumonia and uninfected controls (p<0.005). Plasma levels of inflammatory biomarkers (fractalkine, procalcitonin, pentraxin-3 and suppression of tumorigenicity-2) were independently associated with mcfDNA levels (adjusted p<0.05) among all patients with pneumonia. Such host–microbe interactions in the systemic circulation of patients with severe pneumonia warrant further large-scale clinical and mechanistic investigations.

scholarly journals Multiplex PCR for direct identification of Campylobacter spp. in human and chicken stools

2007 ◽  
Vol 56 (10) ◽  
pp. 1350-1355 ◽  
Author(s):  
Aisha Al Amri ◽  
Abiola C. Senok ◽  
Abdulrahman Yusuf Ismaeel ◽  
Ali E. Al-Mahmeed ◽  
Giuseppe A. Botta
Keyword(s):  
Multiplex Pcr ◽  
Direct Detection ◽  
False Negative ◽  
Enteric Bacteria ◽  
Enzymic Activity ◽  
Campylobacter Coli ◽  
Biochemical Tests ◽  
Direct Evaluation ◽  
Stool Specimens ◽  
Culture Negative

Differentiation between Campylobacter jejuni and Campylobacter coli is problematic in clinical specimens due to fastidious growth requirements and limited biochemical tests. This study describes a rapid, multiplex PCR protocol for the direct detection and differentiation of C. jejuni and C. coli in stools. An evaluation was carried out of this multiplex protocol based on the detection of cadF (genus specific), and hipO (C. jejuni) and asp (C. coli) genes, using stool from patients with Campylobacter enteritis and chicken. Protocol sensitivity was assessed and specificity determined using a panel of enteric bacteria, and evaluation of 30 diarrhoeic stool specimens culture negative for Campylobacter. Of the 114 specimens (54 human and 60 chicken) evaluated by the protocol, 70 (61.4 %) were identified as C. jejuni, 35 (30.7 %) as C. coli and 9 (7.9 %) as a mixed infection/colonization with both species. All mixed infections were identified as C. jejuni by culture. Among the stool specimens that were culture negative for Campylobacter, two (6.7 %) were C. jejuni positive by multiplex PCR. The protocol sensitivity limit was 0.015–0.016 ng C. jejuni and C. coli DNA μl−1 in the specimen. There was no cross-reaction with the reference strains assessed. Comparison of hippurate test and multiplex PCR demonstrated 17 isolates with false-positive hippurate enzymic activity and 7 with false-negative activity. This rapid protocol (turnaround time 6 h) is highly sensitive and specific for direct evaluation of stool for these pathogens. It has significant application for routine clinical diagnostic and epidemiological purposes.

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