Abstract:
:Combination therapy can successfully suppress human immunodeficiency virus (HIV) replication in patients but selects for drug resistance, requiring subsequent resistance-guided therapeutic changes. This report describes the development and validation of a novel assay that offers a uniform method to measure susceptibility to all clinically approved HIV type 1 (HIV-1) drugs targeting reverse transcriptase (RT), protease (PR), integrase (IN), and viral entry. It is an assay in which the antiviral effect on infection within a single replication cycle is measured in triply transfected U87.CD4.CXCR4.CCR5 cells, based on homologous recombination between patient-derived amplicons and molecular proviral clones tagged with the enhanced green fluorescent protein (EGFP) reporter gene and from which certain viral genomic regions are removed. The deletions stretch from p17 codon 7 to PR codon 98 in pNL4.3-DeltagagPR-EGFP, from PR codons 1 to 99 in pNL4.3-DeltaPR-EGFP, from RT codons 1 to 560 in pNL4.3-DeltaRT-EGFP, from IN codons 1 to 288 in pNL4.3-DeltaIN-EGFP, and from gp120 codon 34 to gp41 codon 237 in pNL4.3-Deltaenv-EGFP. The optimized experimental conditions enable the investigation of patient samples regardless of viral subtype or coreceptor use. The extraction and amplification success rate for a set of clinical samples belonging to a broad range of HIV-1 group M genetic forms (A-J, CRF01-03, CRF05, and CRF12-13) and displaying a viral load range of 200 to >500,000 RNA copies/ml was 97%. The drug susceptibility measurements, based on discrimination between infected and noninfected cells on a single-cell level by flow cytometry, were reproducible, with coefficients of variation for resistance ranging from 7% to 31%, and were consistent with scientific literature in terms of magnitude and specificity.
journal_name
J Clin Microbioljournal_title
Journal of clinical microbiologyauthors
Covens K,Dekeersmaeker N,Schrooten Y,Weber J,Schols D,Quiñones-Mateu ME,Vandamme AM,Van Laethem Kdoi
10.1128/JCM.01739-08subject
Has Abstractpub_date
2009-07-01 00:00:00pages
2232-42issue
7eissn
0095-1137issn
1098-660Xpii
JCM.01739-08journal_volume
47pub_type
杂志文章abstract::We describe a simple method for the rapid detection of bovine viral diarrhea virus (BVDV) that uses a one-tube reverse transcription PCR (RT-PCR) and total RNA extracted directly from a variety of bovine specimens, including whole blood and tissues. Reagents for both RT and PCR were combined in a one-tube, single-buff...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.33.2.287-291.1995
更新日期:1995-02-01 00:00:00
abstract::We conducted a Linear Array test/retest analysis using cytologic specimens from 198 women. A total of 67.2% of samples had the same human papillomavirus (HPV) types detected in both tests (type-specific positive agreement was 83.3% overall [Kappa = 0.9] and 86.8% for carcinogenic types [Kappa = 0.92]). Discordance was...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.03036-12
更新日期:2013-02-01 00:00:00
abstract::A new immunochromatographic rapid test, Determine HIV-1/2, for the detection of antibodies to human immunodeficiency virus type 1 (HIV-1) and HIV-2 in human whole blood, serum, and plasma was evaluated. Determine HIV-1/2 is a sandwich immunoassay and uses a nitrocellulose strip with a capture site for the patient's re...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.37.2.367-370.1999
更新日期:1999-02-01 00:00:00
abstract::A large pool of refined endotoxin was prepared from Escherichia coli O113 by extraction with hot acqueous phenol. It was characterized chemically and biologically and will be available for a reference standard designated as reference endotoxin EC. ...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:
更新日期:1976-01-01 00:00:00
abstract::A study was conducted to compare the Bac-T-Screen Bacterial Detection Device for Urines (BDD; Marion Laboratories, Kansas City, Mo.) with urine Gram stain as a screen for bacteriuria. We analyzed 631 urine samples with the BDD and compared the results to urine Gram stains and quantitative cultures. A total of 90 (14%)...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.18.3.674-679.1983
更新日期:1983-09-01 00:00:00
abstract::More than 4,000 clinical urine specimens were evaluated with an automated microbial detection/identification system compared to a standarized manual analysis and the routine modalities used in five peer-group laboratories. The comparison indicates that the automated system recognizes the nine groups of significant mic...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.10.2.226-230.1979
更新日期:1979-08-01 00:00:00
abstract::A rapid and sensitive urease test for mycobacteria utilizing a cumulative radiometric technique is described. Definitive results were obtained within a 30-min incubation period. The procedure is simple and economical. Technical time involved is no greater than in conventional procedures. ...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:
更新日期:1977-06-01 00:00:00
abstract::The Binax NOW assay (Binax, Inc., Portland, Maine) and the BD Directigen EZ assay (Becton Dickinson and Company, Sparks, Md.), two new rapid immunoassays for detection of respiratory syncytial virus (RSV), as well as the BD Directigen RSV assay (DRSV) (Becton Dickinson and Company) and direct immunofluorescence staini...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.42.7.2996-2999.2004
更新日期:2004-07-01 00:00:00
abstract::Cell line 293, a continuous line of transformed human embryonic kidney cells, has been recognized for its sensitivity in the isolation of adenoviruses, particularly the fastidious species 40 and 41, from stool specimens. To explore the possibility of using this cell line for the isolation of other viruses from clinica...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.23.4.704-708.1986
更新日期:1986-04-01 00:00:00
abstract::Treatment of wild rats from a known scrub typhus focus in the Philippines with cyclophosphamide did not significantly enhance the isolation rate of Rickettsia tsutsugamushi using mouse inoculation techniques. Similarly, cyclophosphamide treatment of mice inoculated with organs of wild rats did not increase the recover...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:
更新日期:1977-01-01 00:00:00
abstract::This report describes the first isolation of the ameba Balamuthia mandrillaris from an environmental soil sample associated with a fatal case of amebic encephalitis in a northern California child. Isolation of the ameba into culture from autopsied brain tissue confirmed the presence of Balamuthia: In trying to locate ...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/jcm.41.7.3175-3180.2003
更新日期:2003-07-01 00:00:00
abstract::Using PCR-based denaturing gradient gel electrophoresis analyses of oral bacterial samples in 20 mother-child dyads, this study demonstrated a high degree of similarity of bacterial compositions between the mothers and their children; the two may share as much as 94% of their oral bacterial spectra, including cariogen...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.00771-07
更新日期:2007-09-01 00:00:00
abstract::Recent Lassa virus strains from Nigeria were completely or partially sequenced. Phylogenetic analysis revealed the predominance of lineage II and III strains, the existence of a previously undescribed (sub)lineage in Nigeria, and the directional spread of virus in the southern part of the country. The Bayesian analysi...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.01891-10
更新日期:2011-03-01 00:00:00
abstract::During the 2009 H1N1 influenza A outbreak, 773 children were tested for influenza by direct fluorescent-antibody testing with PCR confirmation. Direct fluorescent-antibody testing has a specificity of 99.6% but a sensitivity of only 65.0%. Physicians should recognize diagnostic limitations of direct fluorescent-antibo...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.05255-11
更新日期:2011-10-01 00:00:00
abstract::The 1896 precore (PC) mutation is the most frequent cause of hepatitis B virus e-antigen (HBeAg)-negative chronic hepatitis B virus (HBV) infection. Detection of the 1896 PC mutation has application in studies monitoring antiviral therapy and the natural history of the disease. Identification of this mutation is usual...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.43.1.254-258.2005
更新日期:2005-01-01 00:00:00
abstract::The relationship of coagulase-negative staphylococcal slime production and adherence with the development and outcome of bloodstream infections in two Finnish hospitals was evaluated. Analysis of 64 strains from 62 adult septicemias disclosed 34 (53%) adherent slime producers. In comparison, only 142 (29%) of 489 sing...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.28.12.2779-2785.1990
更新日期:1990-12-01 00:00:00
abstract::Coprological examination based on egg detection in stool samples is currently used as the gold standard for the diagnosis of human fascioliasis. However, this method is not effective during the acute phase of the disease and has poor sensitivity during the chronic phase. Serodiagnosis has become an excellent alternati...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.02875-13
更新日期:2014-03-01 00:00:00
abstract::Bordetella pertussis is a Gram-negative bacterium that causes respiratory infections in humans. Ongoing molecular surveillance of B. pertussis acellular vaccine (aP) antigens is critical for understanding the interaction between evolutionary pressures, disease pathogenesis, and vaccine effectiveness. Methods currently...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.02436-16
更新日期:2017-05-01 00:00:00
abstract::The polymerase chain reaction (PCR) has many potential applications in the field of nucleic acid diagnostics. In particular, it has been successfully applied to the detection of pathogens present in low copy numbers such as the human immunodeficiency virus type 1. Here we describe a time-resolved fluorescence-based hy...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.29.4.798-804.1991
更新日期:1991-04-01 00:00:00
abstract::A total of 201 Mycoplasma pneumoniae clinical isolates from Beijing, China, isolated from 2008 to 2011, were clustered into 16 multiple-locus variable-number tandem-repeat analysis (MLVA) types, of which 6 new MLVA types have never been reported previously. Type 1 isolates based on p1 gene genotyping were mainly MLVA ...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.02567-12
更新日期:2013-02-01 00:00:00
abstract::A new, rapid method for the detection of human immunodeficiency virus type 1 (HIV-1) antibody by time-resolved fluoroimmunoassay (TR-FIA) was developed. In this assay format, microtitration strips were coated with a recombinant fusion protein, and the same protein was labeled with europium and added into the wells sim...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.28.9.2022-2029.1990
更新日期:1990-09-01 00:00:00
abstract::We have evaluated the diagnostic utility of eleven Toxoplasma gondii recombinant antigens (P22 [SAG2], P24 [GRA1], P25, P28 [GRA2], P29 [GRA7], P30 [SAG1], P35, P41 [GRA4], P54 [ROP2], P66 [ROP1], and P68) in immunoglobulin G (IgG) and IgM recombinant enzyme-linked immunosorbent assays (Rec-ELISAs). Following an initi...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.38.3.1144-1150.2000
更新日期:2000-03-01 00:00:00
abstract::We have compared the sensitivity and specificity of quantitative mycobacterial culture against results obtained by using the polymerase chain reaction for the detection of DNA from organisms of the Mycobacterium tuberculosis complex in 82 clinical specimens from patients suspected of having tuberculosis. Two amplifica...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.29.4.712-717.1991
更新日期:1991-04-01 00:00:00
abstract::The use of rapid-cycle PCR combined with colorimetric microwell hybridization for detecting Bordetella pertussis was investigated. Rapid cycling was performed with an air thermocycler (model 1605; Idaho Technology, Idaho Falls, Idaho). Although the instrument was originally designed to be used with capillary tubes, an...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.34.6.1355-1358.1996
更新日期:1996-06-01 00:00:00
abstract::A direct and an indirect enzyme immunoassay (EIA) were evaluated against a standard of electron microscopy after direct ultracentrifugation of the specimen for their performances in detecting rotaviruses. The indirect EIA had variable background activity which influenced test specificity. The indirect EIA control (tes...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.16.1.53-59.1982
更新日期:1982-07-01 00:00:00
abstract::Typical and atypical Campylobacter strains were isolated from the colons of cattle and swine by techniques that enabled the selective pressures of antibiotics to be avoided. Some cephalothin-susceptible strains and a strain with an indeterminate hippurate reaction were classified as Campylobacter coli by DNA homology ...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.26.3.518-523.1988
更新日期:1988-03-01 00:00:00
abstract::Analysis of the electrophoretic profiles of the outer membrane proteins could be used to differentiate Campylobacter jejuni (16 strains) from Campylobacter coli (10 strains). This observation was confirmed by the study of DNA homology obtained by a quantitative filter hybridization method. The hippurate hydrolysis tes...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.27.5.1072-1076.1989
更新日期:1989-05-01 00:00:00
abstract::We have developed and validated a nested real-time PCR (NRT-PCR) for the genotyping of Chlamydia trachomatis and used it specifically for the typing of either eight genovars from D to K or three genovars of lymphogranuloma venereum (LGV). The 11 probes used in the NRT-PCR correctly identified the DNA from D to K and L...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.00451-07
更新日期:2007-08-01 00:00:00
abstract::Nocardia cyriacigeorgica is a recently characterized species within the genus of Nocardia. We report a brain abscess, following a primary pulmonary colonization, due to this species in a human immunodeficiency virus-infected patient. This case confirms that isolation of Nocardia in sputum is associated with a high ris...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.43.9.4895-4897.2005
更新日期:2005-09-01 00:00:00
abstract::The clinical importance of nondiphtherial Corynebacterium, a ubiquitous member of the normal human microflora of the skin and mucous membrane, for ocular surface infections has been recognized recently. We performed an antimicrobial susceptibility test with Etest strips for three fluoroquinolones (ciprofloxacin, norfl...
journal_title:Journal of clinical microbiology
pub_type: 杂志文章
doi:10.1128/JCM.01741-07
更新日期:2008-02-01 00:00:00