Publications by authors named "David H Persing"

58 Publications

Blood-based host biomarker diagnostics in active case finding for pulmonary tuberculosis: A diagnostic case-control study.

EClinicalMedicine 2021 Mar 6;33:100776. Epub 2021 Mar 6.

Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, CA, USA.

Background: There is a need to identify scalable tuberculosis screening strategies among high burden populations. The WHO has identified a non-sputum-based triage test as a development priority.

Methods: We performed a diagnostic case-control study of point-of-care C-reactive protein (CRP) and Prototype-Xpert-MTB-Host-Response (Xpert-MTB-HR) assays in the context of a mass screening program for tuberculosis in two prisons in Brazil. All incarcerated individuals irrespective of symptoms were screened by sputum Xpert MTB/RIF and sputum culture. Among consecutive, Xpert MTB/RIF or culture-confirmed cases and Xpert MTB/RIF and culture-negative controls, CRP was quantified in serum by a point-of-care assay (iChroma-II) and a 3-gene expression score was quantified from whole blood using the Xpert-MTB-HR cartridge. We evaluated receiver operating characteristic area under the curve (AUC) and assessed specificity at 90% sensitivity and sensitivity at 70% specificity, consistent with WHO target product profile (TPP) benchmarks.

Findings: Two hundred controls (no TB) and 100 culture- or Xpert MTB/RIF-positive tuberculosis cases were included. Half of tuberculosis cases and 11% of controls reported any tuberculosis symptoms. AUC for CRP was 0·79 (95% CI: 0·73-0·84) and for Xpert-MTB-HR was 0·84 (95% CI: 0·79-0·89). At 90% sensitivity, Xpert-MTB-HR had significantly higher specificity (53·0%, 95% CI: 45·0-69·0%) than CRP (28·1%, 95% CI: 20·2-41·8%) ( = 0·003), both well below the TPP benchmark of 70%. Among individuals with medium or high sputum Xpert MTB/RIF semi-quantitative load, sensitivity (at 70% specificity) of CRP (90·3%, 95% CI: 74·2-98·0) and Xpert-MTB-HR (96·8%, 95% CI: 83·3-99·9%) was higher.

Interpretation: For active case finding in this high tuberculosis-burden setting, CRP and Xpert-MTB-HR did not meet TPP benchmarks for a triage test. However, Xpert-MTB-HR was highly sensitive in detecting individuals with medium or high sputum bacillary burden.

Funding: National Institutes of Health (R01 AI130058 and R01 AI149620) and Brazilian National Council for Scientific and Technological Development (CNPq-404182/2019-4).
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http://dx.doi.org/10.1016/j.eclinm.2021.100776DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8020164PMC
March 2021

Multicenter Evaluation of the Cepheid Xpert Xpress SARS-CoV-2/Flu/RSV Test.

J Clin Microbiol 2021 02 18;59(3). Epub 2021 Feb 18.

Cepheid, Sunnyvale, California, USA

With the approach of respiratory virus season in the Northern Hemisphere, clinical microbiology and public health laboratories will need rapid diagnostic assays to distinguish severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from influenza virus and respiratory syncytial virus (RSV) infections for diagnosis and surveillance. In this study, the clinical performance of the Xpert Xpress SARS-CoV-2/Flu/RSV test (Cepheid, Sunnyvale, CA, USA) for nasopharyngeal swab specimens was evaluated in four centers: Johns Hopkins Medical Microbiology Laboratory, Northwell Health Laboratories, NYC Public Health Laboratory, and Los Angeles County/University of Southern California (LAC+USC) Medical Center. A total of 319 nasopharyngeal swab specimens, positive for SARS-CoV-2 ( = 75), influenza A virus ( = 65), influenza B virus ( = 50), or RSV ( = 38) or negative ( = 91) by the standard-of-care nucleic acid amplification tests at each site, were tested using the Cepheid panel test. The overall positive percent agreement for the SARS-CoV-2 target was 98.7% ( = 74/75), and the negative agreement was 100% ( = 91), with all other analytes showing 100% total agreement ( = 153). Standard-of-care tests to which the Cepheid panel was compared included the Cepheid Xpert Xpress SARS-CoV-2, Cepheid Xpert Xpress Flu/RSV, GenMark ePlex respiratory panel, BioFire respiratory panel 2.1 and v1.7, DiaSorin Simplexa COVID-19 Direct, and Hologic Panther Fusion SARS-CoV-2 assays. The Xpert Xpress SARS-CoV-2/Flu/RSV test showed high sensitivity and accuracy for all analytes included in the test. This test will provide a valuable clinical diagnostic and public health solution for detecting and differentiating SARS-CoV-2, influenza A and B virus, and RSV infections during the current respiratory virus season.
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http://dx.doi.org/10.1128/JCM.02955-20DOI Listing
February 2021

Diagnostic Accuracy Study of a Novel Blood-Based Assay for Identification of Tuberculosis in People Living with HIV.

J Clin Microbiol 2021 Feb 18;59(3). Epub 2021 Feb 18.

FIND, Geneva, Switzerland.

A nonsputum triage test to rule out tuberculosis (TB) disease is a WHO high-priority diagnostic, and a combinatory score based on a 3-gene host signature has shown promise in discriminating TB from other illnesses. We evaluated the accuracy of an early-prototype cartridge assay ("Xpert MTB Host Response" or Xpert-MTB-HR-Prototype) of this 3-gene signature on biobanked blood samples from people living with HIV (PLHIV) against a comprehensive microbiological reference standard (CMRS) and against Xpert MTB/RIF on the first sputum sample alone. We depict results based on performance targets set by the WHO in comparison with a laboratory-based C-reactive protein (CRP) assay. Of 201 patients included, 67 were culture positive for The areas under the concentration-time curve (AUCs) for Xpert-MTB-HR-Prototype were 0.89 (confidence interval [CI], 0.83 to 0.94) against the CMRS and 0.94 (CI, 0.89 to 0.98) against Xpert MTB/RIF. Considering Xpert-MTB-HR-Prototype as a triage test (at the nearest upper value of sensitivity to 90%), specificities were 55.8% (CI, 47.2 to 64.1%) compared to the CMRS and 85.9% (CI, 79.3 to 90.7%) compared to Xpert MTB/RIF as confirmatory tests. Considering Xpert-MTB-HR-Prototype as a stand-alone diagnostic test, at a specificity near 95%, the test achieved a sensitivity of 65.7% (CI, 53.7 to 75.9%), while the CRP assay achieved a sensitivity of only 13.6% (CI, 7.3 to 23.4%). In this first accuracy study of a prototype blood-based host marker assay, we show the possible value of the assay for triage and diagnosis in PLHIV.
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http://dx.doi.org/10.1128/JCM.01643-20DOI Listing
February 2021

Real-time Screening of Specimen Pools for Coronavirus Disease 2019 (COVID-19) Infection at Sanya Airport, Hainan Island, China.

Clin Infect Dis 2020 Sep 30. Epub 2020 Sep 30.

Department of Tropical Medicine and Infectious Diseases, Hainan Hospital, The Chinese PLA General Hospital, Sanya, Hainan, China.

A 10:1 pooled test strategy on-site at an airport of China was pursued, resulting in increased test throughput, limited use of reagents, and increased testing efficiency without loss of sensitivity. This testing approach has the potential to reduce the need for contact tracing when the results are delivered first time.
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http://dx.doi.org/10.1093/cid/ciaa1074DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7543279PMC
September 2020

The Xpert MTB/RIF Ultra assay detects Mycobacterium tuberculosis complex DNA in white rhinoceros (Ceratotherium simum) and African elephants (Loxodonta africana).

Sci Rep 2020 09 2;10(1):14482. Epub 2020 Sep 2.

DSI-NRF Centre of Excellence for Biomedical Tuberculosis Research, South African Medical Research Council Centre for Tuberculosis Research, Division of Molecular Biology and Human Genetics, Faculty of Medicine and Health Sciences, Stellenbosch University, PO Box 241, Cape Town, 8000, South Africa.

The study describes the novel use of the Xpert MTB/RIF Ultra assay for detection of Mycobacterium tuberculosis complex (MTBC) DNA in samples from white rhinoceros (Ceratotherium simum) and African elephants (Loxodonta africana). Culture negative respiratory sample matrices were spiked to determine if the Ultra could detect MTBC DNA in rhinoceros and elephant samples. Rhinoceros bronchial alveolar lavage fluid (BALF) was found to have an inhibitory effect on the Ultra. In this study, the limit of detection (LOD) of M. tuberculosis H37Rv in all spiked animal samples were 2 CFU/ml compared to 15.6 CFU/ml for humans, while the LOD for M. bovis SB0121 was 30 CFU/ml compared to 143.4 CFU/ml for M. bovis BCG in humans. Screening was performed on stored tissue and respiratory samples from known MTBC-infected animals and MTBC DNA was detected in 92% of samples collected from six rhinoceros and two elephants. Conversely, 83% of culture-negative tissue and respiratory samples from uninfected animals tested negative on the Ultra. In conclusion, the Ultra assay appears to be a sensitive and rapid diagnostic test for the detection of MTBC DNA from tissue and respiratory samples collected from African elephants and rhinoceros. Furthermore, the Ultra assay could provide a new tool for the detection of MTBC in various sample types from other wildlife species.
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http://dx.doi.org/10.1038/s41598-020-71568-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7468236PMC
September 2020

Multicenter Evaluation of the Cepheid Xpert Xpress SARS-CoV-2 Test.

J Clin Microbiol 2020 Jul 23;58(8). Epub 2020 Jul 23.

Cepheid, Sunnyvale, California, USA.

Nucleic acid amplification tests (NAATs) are the primary means of identifying acute infections caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Accurate and fast test results may permit more efficient use of protective and isolation resources and allow rapid therapeutic interventions. We evaluated the analytical and clinical performance characteristics of the Xpert Xpress SARS-CoV-2 (Xpert) test, a rapid, automated molecular test for SARS-CoV-2. Analytical sensitivity and specificity/interference were assessed with infectious SARS-CoV-2; other infectious coronavirus species, including SARS-CoV; and 85 nasopharyngeal swab specimens positive for other respiratory viruses, including endemic human coronaviruses (hCoVs). Clinical performance was assessed using 483 remnant upper- and lower-respiratory-tract specimens previously analyzed by standard-of-care (SOC) NAATs. The limit of detection of the Xpert test was 0.01 PFU/ml. Other hCoVs, including Middle East respiratory syndrome coronavirus, were not detected by the Xpert test. SARS-CoV, a closely related species in the subgenus , was detected by a broad-range target (E) but was distinguished from SARS-CoV-2 (SARS-CoV-2-specific N2 target). Compared to SOC NAATs, the positive agreement of the Xpert test was 219/220 (99.5%), and the negative agreement was 250/261 (95.8%). A third tie-breaker NAAT resolved all but three of the discordant results in favor the Xpert test. The Xpert test provided sensitive and accurate detection of SARS-CoV-2 in a variety of upper- and lower-respiratory-tract specimens. The high sensitivity and short time to results of approximately 45 min may impact patient management.
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http://dx.doi.org/10.1128/JCM.00926-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7383535PMC
July 2020

Laboratory Diagnosis of COVID-19: Current Issues and Challenges.

J Clin Microbiol 2020 05 26;58(6). Epub 2020 May 26.

Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee, USA.

The COVID-19 outbreak has had a major impact on clinical microbiology laboratories in the past several months. This commentary covers current issues and challenges for the laboratory diagnosis of infections caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In the preanalytical stage, collecting the proper respiratory tract specimen at the right time from the right anatomic site is essential for a prompt and accurate molecular diagnosis of COVID-19. Appropriate measures are required to keep laboratory staff safe while producing reliable test results. In the analytic stage, real-time reverse transcription-PCR (RT-PCR) assays remain the molecular test of choice for the etiologic diagnosis of SARS-CoV-2 infection while antibody-based techniques are being introduced as supplemental tools. In the postanalytical stage, testing results should be carefully interpreted using both molecular and serological findings. Finally, random-access, integrated devices available at the point of care with scalable capacities will facilitate the rapid and accurate diagnosis and monitoring of SARS-CoV-2 infections and greatly assist in the control of this outbreak.
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http://dx.doi.org/10.1128/JCM.00512-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7269383PMC
May 2020

mRNA Detection in Anal Cytology: A Feasible Approach for Anal Cancer Screening in Men Who Have Sex with Men Living With HIV.

Diagnostics (Basel) 2019 11 2;9(4). Epub 2019 Nov 2.

Department of Pathology, Hospital Clínic, University of Barcelona, 08036 Barcelona, Spain.

There is growing interest in anal cancer screening strategies. However, cytological/molecular evaluation of anal samples is challenging. We aimed to determine the feasibility of detecting, in anal liquid-based cytologies, the expression of biomarkers involved in the cell cycle disturbance elicited by human papillomavirus (HPV). The accuracy of this approach in the identification of high-grade squamous intraepithelial lesions/anal intraepithelial neoplasia grade2-3 (HSIL/AIN2-3) was also evaluated. 215 anal cytologies from men having sex with men living with human immunodeficiency virus were evaluated. Patients showing concordant cytological and anoscopy-directed biopsy diagnosis were selected: 70 with negative cytology and HPV test, 70 with low-grade SIL (LSIL/AIN1) cytology and biopsy, and 75 with cytology and biopsy of HSIL/AIN2-3. CDKN2A/p16, MKI67 and TOP2A mRNA expression was analyzed. HPV detection was performed with Xpert HPV Assay (Cepheid, Sunnyvale, CA, USA). HSIL/AIN2-3 showed higher expression for the biomarkers than LSIL/AIN1 or negative samples. The specificity for HSIL/AIN2-3 detection for a sensitivity established at 70% was 44.7% (95%confidence interval [CI] 36.5-53.2) for TOP2A and MKI67 and 54.5% (95%CI 46.0-62.8%) for CDKN2A/p16. mRNA detection of cell biomarkers in anal liquid-based cytology is feasible. Further studies are warranted to confirm if strategies based on mRNA detection have any role in anal cancer screening.
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http://dx.doi.org/10.3390/diagnostics9040173DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6963475PMC
November 2019

Guidelines Support the Value of Stand-Alone Nucleic Acid Amplification Tests for () Infection.

J Clin Microbiol 2019 10 24;57(10). Epub 2019 Sep 24.

University of Washington School of Medicine, Seattle, Washington, USA.

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http://dx.doi.org/10.1128/JCM.01079-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6760941PMC
October 2019

Evaluation of the Xpert HCV Viral Load Finger-Stick Point-of-Care Assay.

J Infect Dis 2018 05;217(12):1889-1896

The Kirby Institute, UNSW Sydney, Sydney, Australia.

Point-of-care hepatitis C virus (HCV) RNA testing is advantageous, enabling diagnosis of active infection in a single visit. This study evaluated the sensitivity and specificity of the Xpert HCV Viral Load Finger-Stick assay (Xpert HCV VL FS) for HCV RNA detection (finger-stick) and the Xpert HCV Viral Load assay (plasma) compared with the Abbott RealTime HCV Viral Load assay by venepuncture. Plasma and finger-stick capillary whole-blood samples were collected from participants in an observational cohort in Australia. Of 223 participants enrolled, HCV RNA was detected in 40% of participants (85 of 210) with available Xpert HCV Viral Load testing. Participants receiving HCV therapy were excluded from subsequent analyses (n = 16). Sensitivity of the Xpert HCV Viral Load assay for HCV RNA quantification in plasma collected by venepuncture was 100.0% (95% confidence interval [CI] 96.9%-100.0%) and specificity was 100.0% (95% CI, 94.4%-100.0%). Sensitivity of the Xpert HCV VL FS assay for HCV RNA quantification in samples collected by finger-stick was 100.0% (95% CI, 93.9%-100.0%) and specificity was 100.0% (95% CI, 96.6%-100.0%). The Xpert HCV VL FS test can accurately detect active infection from a finger-stick sample in 1 hour allowing single-visit HCV diagnosis.
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http://dx.doi.org/10.1093/infdis/jiy114DOI Listing
May 2018

Clinical Impact and Cost-effectiveness of Xpert MTB/RIF Testing in Hospitalized Patients With Presumptive Pulmonary Tuberculosis in the United States.

Clin Infect Dis 2017 02;64(4):482-489

Harborview Medical Center.

Background: Microscopic examination of acid-fast-stained sputum smears is the current standard of care in the United States to determine airborne infection isolation (AII) of inpatients with presumptive pulmonary tuberculosis (PTB). However, nucleic acid amplification testing (NAAT) with the Xpert MTB/RIF assay (Xpert) may be more efficient and less costly.

Methods: This prospective observational cohort study enrolled a consecutive sample of 318 AII-eligible inpatients from a public hospital in Seattle, Washington, from March 2012 to October 2013. Sputum samples were collected from each inpatient and analyzed using smear microscopy, culture, drug susceptibility testing, and NAAT. The performance, clinical utility (AII duration and survival), and cost-effectiveness from an institutional perspective were compared for 5 testing strategies.

Results: Among the 318 admissions with presumptive PTB, 20 (6.3%) were culture-positive for Mycobacterium tuberculosis. The sensitivity of 1 Xpert, 2 Xperts, 2 smears, or 3 smears compared to culture was 0.85 (95% confidence interval [CI], .61–.96), 0.95 (95% CI, .73–1.0), 0.70 (95% CI, .46–.88), and 0.80 (95% CI, .56–.93), respectively. A cost-effectiveness analysis of the study results demonstrated that an Xpert test on 1 unconcentrated sputum sample (assuming equivalent results for unconcentrated and concentrated sputum samples) is the most cost-effective strategy (99.9% preferred at willingness-to-pay of US$50000) and on average would save 51.5 patient-hours in AII and up to $11466 relative to microscopy without a compromise in sensitivity.

Conclusions: In hospitalized patients with presumptive PTB in a low-burden setting, NAAT can reduce AII and is comparably sensitive, more specific, and more cost-effective than smear microscopy.
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http://dx.doi.org/10.1093/cid/ciw803DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5399932PMC
February 2017

Analytical Performance Characteristics of the Cepheid GeneXpert Ebola Assay for the Detection of Ebola Virus.

PLoS One 2015 12;10(11):e0142216. Epub 2015 Nov 12.

Cepheid, Sunnyvale, California, United States of America.

Background: The recently developed Xpert® Ebola Assay is a novel nucleic acid amplification test for simplified detection of Ebola virus (EBOV) in whole blood and buccal swab samples. The assay targets sequences in two EBOV genes, lowering the risk for new variants to escape detection in the test. The objective of this report is to present analytical characteristics of the Xpert® Ebola Assay on whole blood samples.

Methods And Findings: This study evaluated the assay's analytical sensitivity, analytical specificity, inclusivity and exclusivity performance in whole blood specimens. EBOV RNA, inactivated EBOV, and infectious EBOV were used as targets. The dynamic range of the assay, the inactivation of virus, and specimen stability were also evaluated. The lower limit of detection (LoD) for the assay using inactivated virus was estimated to be 73 copies/mL (95% CI: 51-97 copies/mL). The LoD for infectious virus was estimated to be 1 plaque-forming unit/mL, and for RNA to be 232 copies/mL (95% CI 163-302 copies/mL). The assay correctly identified five different Ebola viruses, Yambuku-Mayinga, Makona-C07, Yambuku-Ecran, Gabon-Ilembe, and Kikwit-956210, and correctly excluded all non-EBOV isolates tested. The conditions used by Xpert® Ebola for inactivation of infectious virus reduced EBOV titer by ≥6 logs.

Conclusion: In summary, we found the Xpert® Ebola Assay to have high analytical sensitivity and specificity for the detection of EBOV in whole blood. It offers ease of use, fast turnaround time, and remote monitoring. The test has an efficient viral inactivation protocol, fulfills inclusivity and exclusivity criteria, and has specimen stability characteristics consistent with the need for decentralized testing. The simplicity of the assay should enable testing in a wide variety of laboratory settings, including remote laboratories that are not capable of performing highly complex nucleic acid amplification tests, and during outbreaks where time to detection is critical.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0142216PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4643052PMC
June 2016

Latent tuberculosis: interferon and beyond?

Authors:
David H Persing

J Mol Diagn 2015 Jan;17(1):2-3

Cepheid, Sunnyvale, California. Electronic address:

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http://dx.doi.org/10.1016/j.jmoldx.2014.08.004DOI Listing
January 2015

mRNA biomarker detection in liquid-based cytology: a new approach in the prevention of cervical cancer.

Mod Pathol 2015 Feb 5;28(2):312-20. Epub 2014 Sep 5.

Faculty of Medicine, Department of Pathology, CRESIB (Centre de Recerca en Salut Internacional de Barcelona), Hospital Clínic, University of Barcelona, Barcelona, Spain.

Several high-risk human papillomavirus (HPV)-induced cell biomarkers have been proposed as possible candidates to identify patients harboring high-grade squamous intraepithelial lesions (HSILs) of the uterine cervix. We aimed to determine the feasibility of the detection of the mRNA of six biomarkers in cervical smear specimens obtained by liquid-based cytology and to evaluate whether this approach might be useful in the identification of patients with HSIL. One-hundred and twenty three women referred to colposcopy in the Hospital Clinic of Barcelona were included in the study. After a thorough study, including Pap test, high-risk HPV testing (Hybrid Capture 2 test), and colposcopy with directed biopsy and/or endocervical curettage, 48 patients were diagnosed with HSIL, whereas 75 were classified as negative (n=28), or harboring low-grade SIL (n=47). CDKN2A/p16, BIRC5, MMP9, TOP2A, MCM5, and MKI67 mRNA expression was analyzed by reverse transcription quantitative polymerase chain reaction in liquid-based cytology after the Pap test and Hybrid Capture 2 performance. The tissue expression of these biomarkers was analyzed by immunohistochemistry in the biopsy material. One-hundred and thirteen out of 123 (92%) liquid-based cytology yielded adequate material for mRNA analysis. TOP2A was the most sensitive (97%) biomarker for the detection of HSIL and CDKN2A/p16 the most specific (78%). The combination of TOP2A and CDKN2A/p16 showed a sensitivity of 96% (95% confidence interval (CI): 88-99) and a specificity of 71% (95% CI: 55-82). In the immunohistochemistry analysis, all biomarkers showed a high sensitivity but low specificity for HSIL, except CDKN2A/p16 which had a sensitivity of 100% and a specificity of 63%. The combination of TOP2A and CDKN2A/p16 showed a sensitivity of 100% (95% CI: 91-100) and a specificity of 43% (95% CI: 32-55). The detection of mRNA of cell biomarkers in liquid-based cytology material is feasible. The combination TOP2A and CDKN2A/p16 has a good balance between sensitivity and specificity for the detection of women with HSIL.
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http://dx.doi.org/10.1038/modpathol.2014.106DOI Listing
February 2015

Strain types and antimicrobial resistance patterns of Clostridium difficile isolates from the United States, 2011 to 2013.

Antimicrob Agents Chemother 2014 Jul 21;58(7):4214-8. Epub 2014 Apr 21.

Cepheid, Sunnyvale, California, USA.

We determined the PCR ribotypes and antimicrobial susceptibility patterns of 508 toxigenic Clostridium difficile isolates collected between 2011 and 2013 from 32 U.S. hospitals. Of the 29 PCR ribotypes identified, the 027 strain type was the most common (28.1%), although the rates varied by geographic region. Ribotype 014/020 isolates appear to be emerging. Clindamycin and moxifloxacin resistances (36.8% and 35.8%, respectively) were the most frequent resistance phenotypes observed. Reduced susceptibility to vancomycin was observed in 39.1% of 027 isolates.
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http://dx.doi.org/10.1128/AAC.02775-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4068552PMC
July 2014

Genome-wide sequencing of cellular microRNAs identifies a combinatorial expression signature diagnostic of sepsis.

PLoS One 2013 16;8(10):e75918. Epub 2013 Oct 16.

Department of Experimental Immunobiology, King's College London, London, United Kingdom ; NIHR Biomedical Research Centre, Guy's and St Thomas' NHS Foundation Trust and King's College London, London, United Kingdom.

Rationale: Sepsis is a common cause of death in the intensive care unit with mortality up to 70% when accompanied by multiple organ dysfunction. Rapid diagnosis and the institution of appropriate antibiotic therapy and pressor support are therefore critical for survival. MicroRNAs are small non-coding RNAs that play an important role in the regulation of numerous cellular processes, including inflammation and immunity.

Objectives: We hypothesized changes in expression of microRNAs during sepsis may be of diagnostic value in the intensive care unit (ICU).

Methods: Massively parallel sequencing of microRNAs was utilised for screening microRNA candidates. Putative microRNAs were validated using quantitative real-time PCR (qRT-PCR). This study includes data from both a training cohort (UK) and an independent validation cohort (Sweden). A linear discriminant statistical model was employed to construct a diagnostic microRNA signature.

Results: A panel of known and novel microRNAs were detectable in the blood of patients with sepsis. After qRT-PCR validation, microRNA miR-150 and miR-4772-5p-iso were able to discriminate between patients who have systemic inflammatory response syndrome and patients with sepsis. This finding was also validated in independent cohort with an average diagnostic accuracy of 86%. Fractionating the cellular components of blood reveals miR-4772-5p-iso is expressed differentially in monocytes. Functional experiments using primary human monocytes demonstrate that it expressed in response to TLR ligation.

Conclusions: Taken together, these data provide a novel microRNA signature of sepsis that should allow rapid point-of-care diagnostic assessment of patients on ICU and also provide greater insight into the pathobiology of this severe disease.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0075918PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3797812PMC
July 2014

Detection of colonization by carbapenemase-producing Gram-negative Bacilli in patients by use of the Xpert MDRO assay.

J Clin Microbiol 2013 Nov 4;51(11):3780-7. Epub 2013 Sep 4.

Cepheid, Sunnyvale, California, USA.

Detecting colonization of patients with carbapenemase-producing bacteria can be difficult. This study compared the sensitivity and specificity of a PCR-based method (Xpert MDRO) for detecting blaKPC, blaNDM, and blaVIM carbapenem resistance genes using GeneXpert cartridges to the results of culture with and without a broth enrichment step on 328 rectal, perirectal, and stool samples. The culture method included direct inoculation of a MacConkey agar plate on which a 10-μg meropenem disk was placed and plating on MacConkey agar after overnight enrichment of the sample in MacConkey broth containing 1 μg/ml of meropenem. Forty-three (13.1%) samples were positive by PCR for blaKPC and 11 (3.4%) were positive for blaVIM; none were positive for blaNDM. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the PCR assay for blaKPC were 100%, 99.0%, 93.0%, and 100%, respectively, compared to broth enrichment culture and sequencing of target genes. The sensitivity, specificity, PPV, and NPV of the assay for blaVIM were 100%, 99.4%, 81.8%, and 100%, respectively. Since none of the clinical samples contained organisms with blaNDM, 66 contrived stool samples were prepared at various dilutions using three Klebsiella pneumoniae isolates containing blaNDM. The PCR assay showed 100% positivity at dilutions from 300 to 1,800 CFU/ml and 93.3% at 150 CFU/ml. The Xpert MDRO PCR assay required 2 min of hands-on time and 47 min to complete. Rapid identification of patients colonized with carbapenemase-producing organisms using multiplex PCR may help hospitals to improve infection control activities.
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http://dx.doi.org/10.1128/JCM.01092-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3889767PMC
November 2013

Alternative processing of the U2 small nuclear RNA produces a 19-22nt fragment with relevance for the detection of non-small cell lung cancer in human serum.

PLoS One 2013 20;8(3):e60134. Epub 2013 Mar 20.

Service de Pneumologie, Hôpital Larrey, CHU de Toulouse, Université de Toulouse III, Paul Sabatier, Toulouse, France.

RNU2 exists in two functional forms (RNU2-1 and RNU2-2) distinguishable by the presence of a unique 4-bases motif. Detailed investigation of datasets obtained from deep sequencing of five human lung primary tumors revealed that both forms express at a high rate a 19-22nt fragment (miR-U2-1 and -2) from its 3' region and contains the 4-bases motif. Deep sequencing of independent pools of serum samples from healthy donors and lung cancer patients revealed that miR-U2-1 and -2 are pervasively processed in lung tissue by means of endonucleolytic cleavages and stably exported to the blood. Then, microarrays hybridization experiments of matched normal/tumor samples revealed a significant over-expression of miR-U2-1 in 14 of 18 lung primary tumors. Subsequently, qRT-PCR of miR-U2-1 using serum from 62 lung cancer patients and 96 various controls demonstrated that its expression levels identify lung cancer patients with 79% sensitivity and 80% specificity. miR-U2-1 expression correlated with the presence or absence of lung cancer in patients with chronic obstructive pulmonary disease (COPD), other diseases of the lung - not cancer, and in healthy controls. These data suggest that RNU2-1 is a new bi-functional ncRNA that produces a 19-22nt fragment which may be useful in detecting lung cancer non-invasively in high risk patients.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0060134PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3603938PMC
September 2013

Antimicrobial-resistant strains of Clostridium difficile from North America.

Antimicrob Agents Chemother 2012 Jun 12;56(6):2929-32. Epub 2012 Mar 12.

Cepheid, Sunnyvale, California, USA.

A total of 316 toxigenic Clostridium difficile clinical isolates of known PCR ribotypes from patients in North America were screened for resistance to clindamycin, metronidazole, moxifloxacin, and rifampin. Clindamycin resistance was observed among 16 different ribotypes, with ribotypes 017, 053, and 078 showing the highest proportions of resistance. All isolates were susceptible to metronidazole. Moxifloxacin resistance was present in >90% of PCR-ribotype 027 and 053 isolates but was less common among other ribotypes. Only 7.9% of the C. difficile isolates were resistant to rifampin. Multidrug resistance (clindamycin, moxifloxacin, and rifampin) was present in 27.5% of PCR-ribotype 027 strains but was rare in other ribotypes. These results suggest that antimicrobial resistance in North American isolates of C. difficile varies by strain type and parallels rates of resistance reported from Europe and the Far East.
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http://dx.doi.org/10.1128/AAC.00220-12DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3370774PMC
June 2012

Characterization of nasal and blood culture isolates of methicillin-resistant Staphylococcus aureus from patients in United States Hospitals.

Antimicrob Agents Chemother 2012 Mar 12;56(3):1324-30. Epub 2011 Dec 12.

Cepheid, Sunnyvale, California, USA.

A total of 299 nares and 194 blood isolates of methicillin-resistant Staphylococcus aureus (MRSA), each recovered from a unique patient, were collected from 23 U.S. hospitals from May 2009 to March 2010. All isolates underwent spa and staphylococcal cassette chromosome mec element (SCCmec) typing and antimicrobial susceptibility testing; a subset of 84 isolates was typed by pulsed-field gel electrophoresis (PFGE) using SmaI. Seventy-six spa types were observed among the isolates. Overall, for nasal isolates, spa type t002-SCCmec type II (USA100) was the most common strain type (37% of isolates), while among blood isolates, spa type t008-SCCmec type IV (USA300) was the most common (39%). However, the proportion of all USA100 and USA300 isolates varied by United States census region. Nasal isolates were more resistant to tobramycin and clindamycin than blood isolates (55.9% and 48.8% of isolates versus 36.6% and 39.7%, respectively; for both, P < 0.05). The USA300 isolates were largely resistant to fluoroquinolones. High-level mupirocin resistance was low among all spa types (<5%). SCCmec types III and VIII, which are rare in the United States, were observed along with several unusual PFGE types, including CMRSA9, EMRSA15, and the PFGE profile associated with sequence type 239 (ST239) isolates. Typing data from this convenience sample suggest that in U.S. hospitalized patients, USA100 isolates of multiple spa types, while still common in the nares, have been replaced by USA300 isolates as the predominant MRSA strain type in positive blood cultures.
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http://dx.doi.org/10.1128/AAC.05804-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3294931PMC
March 2012

Laboratory diagnosis of Clostridium difficile infection can molecular amplification methods move us out of uncertainty?

J Mol Diagn 2011 Nov 18;13(6):573-82. Epub 2011 Aug 18.

Cepheid, Sunnyvale, CA, USA.

The laboratory diagnosis of Clostridium difficile infection (CDI) continues to be challenging. Recent guidelines from professional societies in the United States note that enzyme immunoassays for toxins A and B do not have adequate sensitivity to be used alone for detecting CDI, yet the optimal method for diagnosing this infection remains unclear. Nucleic acid amplification tests (NAATs) that target chromosomal toxin genes (usually the toxin B gene, tcdB) show high sensitivity and specificity, provide rapid results, and are amenable to both batch and on-demand testing, but these tests were not universally recommended for routine use in the recent guidelines. Rather, two-step algorithms that use glutamate dehydrogenase (GDH) assays to screen for C. difficile in stool specimens, followed by either direct cytotoxin testing or culture to identify toxin-producing C. difficile isolates, were recommended in one guideline and either GDH algorithms or NAATs were recommended in another guideline. Unfortunately, neither culture nor direct cytotoxin testing is widely available. In addition, this two-step approach requires 48 to 92 hours to complete, which may delay the initiation of therapy and critical infection control measures. Recent studies also show the sensitivity of several GDH assays to be <90%. This review considers the role of NAATs for diagnosing CDI and explores their potential advantages over two-step algorithms, including shorter time to results, while providing comparable, if not superior, accuracy.
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http://dx.doi.org/10.1016/j.jmoldx.2011.06.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3194048PMC
November 2011

A multisite assessment of the quantitative capabilities of the Xpert MTB/RIF assay.

Am J Respir Crit Care Med 2011 Nov;184(9):1076-84

Division of Infectious Disease, Department of Medicine, University of Medicine and Dentistry of New Jersey, Newark, USA.

Rationale: The Xpert MTB/RIF is an automated molecular test for Mycobacterium tuberculosis that estimates bacterial burden by measuring the threshold-cycle (Ct) of its M. tuberculosis-specific real-time polymerase chain reaction. Bacterial burden is an important biomarker for disease severity, infection control risk, and response to therapy.

Objectives: Evaluate bacterial load quantitation by Xpert MTB/RIF compared with conventional quantitative methods.

Methods: Xpert MTB/RIF results were compared with smear-microscopy, semiquantiative solid culture, and time-to-detection in liquid culture for 741 patients and 2,008 samples tested in a multisite clinical trial. An internal control real-time polymerase chain reaction was evaluated for its ability to identify inaccurate quantitative Xpert MTB/RIF results.

Measurements And Main Results: Assays with an internal control Ct greater than 34 were likely to be inaccurately quantitated; this represented 15% of M. tuberculosis-positive tests. Excluding these, decreasing M. tuberculosis Ct was associated with increasing smear microscopy grade for smears of concentrated sputum pellets (r(s) = -0.77) and directly from sputum (r(s) =-0.71). A Ct cutoff of approximately 27.7 best predicted smear-positive status. The association between M. tuberculosis Ct and time-to-detection in liquid culture (r(s) = 0.68) and semiquantitative colony counts (r(s) = -0.56) was weaker than smear. Tests of paired same-patient sputum showed that high viscosity sputum samples contained ×32 more M. tuberculosis than nonviscous samples. Comparisons between the grade of the acid-fast bacilli smear and Xpert MTB/RIF quantitative data across study sites enabled us to identify a site outlier in microscopy.

Conclusions: Xpert MTB/RIF quantitation offers a new, standardized approach to measuring bacterial burden in the sputum of patients with tuberculosis.
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http://dx.doi.org/10.1164/rccm.201103-0536OCDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3208646PMC
November 2011

Activity of ACHN-490 against meticillin-resistant Staphylococcus aureus (MRSA) isolates from patients in US hospitals.

Int J Antimicrob Agents 2011 Oct 5;38(4):352-4. Epub 2011 Aug 5.

Cepheid, 904 Caribbean Drive, Sunnyvale, CA 94089, USA.

The activity of ACHN-490 was evaluated against 493 meticillin-resistant Staphylococcus aureus (MRSA) isolates collected in 2009-2010 from 23 US hospitals. The MIC(50) and MIC(90) values (minimal inhibitory concentrations for 50% and 90% of the organisms, respectively) for ACHN-490 were 1 and 2 μg/mL compared with 8 and 32 μg/mL for amikacin, 0.5 and 1 μg/mL for gentamicin and 2 and >16 μg/mL for tobramycin. The gene encoding the aminoglycoside-modifying enzyme APH(2″)-Ia/AAC(6')-Ie was present in 12% of the subset of 84 isolates examined by polymerase chain reaction (PCR), whilst the gene encoding ANT(4')-Ia was present in 89% of isolates. ACHN-490 activity was not affected by either enzyme.
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http://dx.doi.org/10.1016/j.ijantimicag.2011.05.016DOI Listing
October 2011

Comparison of strain typing results for Clostridium difficile isolates from North America.

J Clin Microbiol 2011 May 9;49(5):1831-7. Epub 2011 Mar 9.

Cepheid, Sunnyvale, California 94089, USA.

Accurate strain typing is critical for understanding the changing epidemiology of Clostridium difficile infections. We typed 350 isolates of toxigenic C. difficile from 2008 to 2009 from seven laboratories in the United States and Canada. Typing was performed by PCR-ribotyping, pulsed-field gel electrophoresis (PFGE), and restriction endonuclease analysis (REA) of whole-cell DNA. The Cepheid Xpert C. difficile test for presumptive identification of 027/NAP1/BI isolates was also tested directly on original stool samples. Of 350 isolates, 244 (70%) were known PCR ribotypes, 224 (68%) were 1 of 8 common REA groups, and 187 (54%) were known PFGE types. Eighty-four isolates typed as 027, NAP1, and BI, and 83 of these were identified as presumptive 027/NAP1/BI by Xpert C. difficile. Eight additional isolates were called presumptive 027/NAP1/BI by Xpert C. difficile, of which three were ribotype 027. Five PCR ribotypes contained multiple REA groups, and three North American pulsed-field (NAP) profiles contained both multiple REA groups and PCR ribotypes. There was modest concordance of results among the three methods for C. difficile strains, including the J strain (ribotype 001 and PFGE NAP2), the toxin A-negative 017 strain (PFGE NAP9 and REA type CF), the 078 animal strain (PFGE NAP7 and REA type BK), and type 106 (PFGE NAP11 and REA type DH). PCR-ribotyping, REA, and PFGE provide different but overlapping patterns of strain clustering. Unlike the other methods, the Xpert C. difficile 027/NAP1/BI assay gave results directly from stool specimens, required only 45 min to complete, but was limited to detection of a single strain type.
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http://dx.doi.org/10.1128/JCM.02446-10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3122689PMC
May 2011

Evaluation of a rapid and completely automated real-time reverse transcriptase PCR assay for diagnosis of enteroviral meningitis.

J Clin Microbiol 2011 Feb 15;49(2):528-33. Epub 2010 Dec 15.

Emory University School of Medicine, Atlanta, Georgia, USA.

Nucleic acid amplification tests (NAATs) for enterovirus RNA in cerebrospinal fluid (CSF) have emerged as the new gold standard for diagnosis of enteroviral meningitis, and their use can improve the management and decrease the costs for caring for children with enteroviral meningitis. The Xpert EV assay (Cepheid, Sunnyvale, CA) is a rapid, fully automated real-time PCR test for the detection of enterovirus RNA that was approved by the U.S. Food and Drug Administration for in vitro diagnostic use in March 2007. In this multicenter trial we established the clinical performance characteristics of the Xpert EV assay in patients presenting with meningitis symptoms relative to clinical truth. Clinical truth for enteroviral meningitis was defined as clinical evidence of meningitis, the absence of another detectable pathogen in CSF, and detection of enterovirus in CSF either by two reference NAATs or by viral culture. A total of 199 prospectively and 235 retrospectively collected specimens were eligible for inclusion in this study. The overall prevalence of enteroviral meningitis was 26.04%. The Xpert EV assay had a sensitivity of 94.69% (90% confidence interval [CI] = 89.79 to 97.66%), specificity of 100% (90% CI = 99.07 to 100%), positive predictive value of 100%, negative predictive value of 98.17, and an accuracy of 98.62% relative to clinical truth. The Xpert EV assay demonstrated a high degree of accuracy for diagnosis of enteroviral meningitis. The simplicity and on-demand capability of the Xpert EV assay should prove to be a valuable adjunct to the evaluation of suspected meningitis cases.
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http://dx.doi.org/10.1128/JCM.01570-10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3043510PMC
February 2011

Rapid molecular detection of tuberculosis and rifampin resistance.

N Engl J Med 2010 Sep 1;363(11):1005-15. Epub 2010 Sep 1.

Foundation for Innovative New Diagnostics, Geneva, Switzerland.

Background: Global control of tuberculosis is hampered by slow, insensitive diagnostic methods, particularly for the detection of drug-resistant forms and in patients with human immunodeficiency virus infection. Early detection is essential to reduce the death rate and interrupt transmission, but the complexity and infrastructure needs of sensitive methods limit their accessibility and effect.

Methods: We assessed the performance of Xpert MTB/RIF, an automated molecular test for Mycobacterium tuberculosis (MTB) and resistance to rifampin (RIF), with fully integrated sample processing in 1730 patients with suspected drug-sensitive or multidrug-resistant pulmonary tuberculosis. Eligible patients in Peru, Azerbaijan, South Africa, and India provided three sputum specimens each. Two specimens were processed with N-acetyl-L-cysteine and sodium hydroxide before microscopy, solid and liquid culture, and the MTB/RIF test, and one specimen was used for direct testing with microscopy and the MTB/RIF test.

Results: Among culture-positive patients, a single, direct MTB/RIF test identified 551 of 561 patients with smear-positive tuberculosis (98.2%) and 124 of 171 with smear-negative tuberculosis (72.5%). The test was specific in 604 of 609 patients without tuberculosis (99.2%). Among patients with smear-negative, culture-positive tuberculosis, the addition of a second MTB/RIF test increased sensitivity by 12.6 percentage points and a third by 5.1 percentage points, to a total of 90.2%. As compared with phenotypic drug-susceptibility testing, MTB/RIF testing correctly identified 200 of 205 patients (97.6%) with rifampin-resistant bacteria and 504 of 514 (98.1%) with rifampin-sensitive bacteria. Sequencing resolved all but two cases in favor of the MTB/RIF assay.

Conclusions: The MTB/RIF test provided sensitive detection of tuberculosis and rifampin resistance directly from untreated sputum in less than 2 hours with minimal hands-on time. (Funded by the Foundation for Innovative New Diagnostics.)
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http://dx.doi.org/10.1056/NEJMoa0907847DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2947799PMC
September 2010

Impact of strain type on detection of toxigenic Clostridium difficile: comparison of molecular diagnostic and enzyme immunoassay approaches.

J Clin Microbiol 2010 Oct 11;48(10):3719-24. Epub 2010 Aug 11.

Cepheid, 904 Caribbean Drive, Sunnyvale, CA 98089, USA.

A multicenter clinical trial assessed the performance of the Cepheid Xpert C. difficile assay on stool specimens collected from patients suspected of having Clostridium difficile infection (CDI). A total of 2,296 unformed stool specimens, collected from seven study sites, were tested by Xpert C. difficile enrichment culture followed by cell culture cytotoxicity testing of the isolates (i.e., toxigenic culture with enrichment) and the study sites' standard C. difficile test methods. The methods included enzyme immunoassay (EIA), direct cytotoxin testing, and two- and three-step algorithms using glutamate dehydrogenase (GDH) screening followed by either EIA or EIA and an in-house PCR assay. All C. difficile strains were typed by PCR-ribotyping. Compared to results for toxigenic culture with enrichment, the sensitivity, specificity, and positive and negative predictive values of the Xpert assay were 93.5, 94.0, 73.0, and 98.8%, respectively. The overall sensitivity of the EIAs compared to that of enrichment culture was 60.0%, and the sensitivity of combined GDH algorithms was 72.9%; both were significantly lower than that of Xpert C. difficile (P < 0.001 and P = 0.03, respectively). The sensitivity of the EIA was significantly lower than that of the Xpert C. difficile assay for detection of ribotypes 002, 027, and 106 (P < 0.0001, P < 0.0001, and P = 0.004, respectively, Fisher's exact test), and the sensitivity of GDH algorithms for ribotypes other than 027 was lower than that for Xpert C. difficile (P < 0.001). The Xpert C. difficile assay is a simple, rapid, and accurate method for detection of toxigenic C. difficile in unformed stool specimens and is minimally affected by strain type compared to EIA and GDH-based methods.
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http://dx.doi.org/10.1128/JCM.00427-10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2953097PMC
October 2010

Rapid detection of Mycobacterium tuberculosis and rifampin resistance by use of on-demand, near-patient technology.

J Clin Microbiol 2010 Jan 28;48(1):229-37. Epub 2009 Oct 28.

Department of Medicine, New Jersey Medical School, University of Medicine and Dentistry, New Jersey, Newark, New Jersey 07103, USA.

Current nucleic acid amplification methods to detect Mycobacterium tuberculosis are complex, labor-intensive, and technically challenging. We developed and performed the first analysis of the Cepheid Gene Xpert System's MTB/RIF assay, an integrated hands-free sputum-processing and real-time PCR system with rapid on-demand, near-patient technology, to simultaneously detect M. tuberculosis and rifampin resistance. Analytic tests of M. tuberculosis DNA demonstrated a limit of detection (LOD) of 4.5 genomes per reaction. Studies using sputum spiked with known numbers of M. tuberculosis CFU predicted a clinical LOD of 131 CFU/ml. Killing studies showed that the assay's buffer decreased M. tuberculosis viability by at least 8 logs, substantially reducing biohazards. Tests of 23 different commonly occurring rifampin resistance mutations demonstrated that all 23 (100%) would be identified as rifampin resistant. An analysis of 20 nontuberculosis mycobacteria species confirmed high assay specificity. A small clinical validation study of 107 clinical sputum samples from suspected tuberculosis cases in Vietnam detected 29/29 (100%) smear-positive culture-positive cases and 33/39 (84.6%) or 38/53 (71.7%) smear-negative culture-positive cases, as determined by growth on solid medium or on both solid and liquid media, respectively. M. tuberculosis was not detected in 25/25 (100%) of the culture-negative samples. A study of 64 smear-positive culture-positive sputa from retreatment tuberculosis cases in Uganda detected 63/64 (98.4%) culture-positive cases and 9/9 (100%) cases of rifampin resistance. Rifampin resistance was excluded in 54/55 (98.2%) susceptible cases. Specificity rose to 100% after correcting for a conventional susceptibility test error. In conclusion, this highly sensitive and simple-to-use system can detect M. tuberculosis directly from sputum in less than 2 h.
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http://dx.doi.org/10.1128/JCM.01463-09DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2812290PMC
January 2010

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

J Clin Microbiol 2009 Sep 15;47(9):3008-10. Epub 2009 Jul 15.

Department of Pathology, Vanderbilt University Medical Center, Nashville, Tennessee, USA.

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.
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http://dx.doi.org/10.1128/JCM.01238-09DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2738114PMC
September 2009