Diagnostic performance of Ion 16S metagenomics kit and Ion Reporter metagenomics workflow for bacterial pathogen detection in culture-negative clinical specimens from sterile sources

2021 ◽  
pp. 115451
Author(s):  
Michael Malczynski ◽  
Alex Zhu ◽  
Teresa Zembower ◽  
Chao Qi
Keyword(s):  
Diagnostic Performance ◽  
Pathogen Detection ◽  
Bacterial Pathogen ◽  
Clinical Specimens ◽  
Culture Negative

scholarly journals Comparative Evaluation of Enteric Bacterial Culture and a Molecular Multiplex Syndromic Panel in Children with Acute Gastroenteritis

2019 ◽  
Vol 57 (6) ◽  
Author(s):  
Thomas Kellner ◽  
Brendon Parsons ◽  
Linda Chui ◽  
Byron M. Berenger ◽  
Jianling Xie ◽  
...  
Keyword(s):  
Pathogen Detection ◽  
Rectal Swab ◽  
Bacterial Pathogen ◽  
Positive Culture ◽  
Content Type ◽  
Negative Culture ◽  
Percent Agreement ◽  
Stool Samples ◽  
Culture Positive ◽  
Culture Negative

ABSTRACTAlthough enteric multianalyte syndromic panels are increasingly employed, direct comparisons with traditional methods and the inclusion of host phenotype correlations are limited. Luminex xTAG gastrointestinal pathogen panel (GPP) and culture results are highly concordant. However, phenotypic and microbiological confirmatory testing raises concerns regarding the accuracy of the GPP, especially forSalmonellaspp. A total of 3,089 children with gastroenteritis submitted stool specimens, rectal swab specimens, and clinical data. The primary outcome was bacterial pathogen detection agreement for shared targets between culture and the Luminex xTAG GPP. Secondary analyses included phenotype assessment, additional testing of GPP-negative/culture-positive isolate suspensions with the GPP, and in-house and commercial confirmatory nucleic acid testing of GPP-positive/culture-negative extracts. The overall percent agreement between technologies was >99% for each pathogen.Salmonellaspp. were detected in specimens from 64 participants: 12 (19%) by culture only, 9 (14%) by GPP only, and 43 (67%) by both techniques. Positive percent agreement forSalmonellaspp. was 78.2% (95% confidence interval [CI], 64.6%, 87.8%). Isolate suspensions from the 12 participants with specimens GPP negative/culture positive forSalmonellatested positive by GPP. Specimens GPP positive/culture negative forSalmonellaoriginated in younger children with less diarrhea and more vomiting. GPP-positive/culture-negative specimen extracts tested positive using additional assays for 0/2Campylobacter-positive specimens, 0/4Escherichia coliO157-positive specimens, 0/9Salmonella-positive specimens, and 2/3Shigella-positive specimens. For both rectal swab and stool samples, the median cycle threshold (CT) values, determined using quantitative PCR, were higher for GPP-negative/culture-positive samples than for GPP-positive/culture-positive samples (for rectal swabs, 36.9 [interquartile range {IQR}, 33.7, 37.1] versus 30.0 [IQR, 26.2, 33.2], respectively [P = 0.002]; for stool samples, 36.9 [IQR, 33.7, 37.1] versus 29.0 [IQR, 24.8, 30.8], respectively [P = 0.001]). GPP and culture have excellent overall agreement; however, for specific pathogens, GPP is less sensitive than culture and, notably, identifies samples false positive forSalmonellaspp.

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.

Abstract 11119: Real-Time Nanopore Sequencing for Bacterial Pneumonia After Out-of-Hospital Cardiac Arrest, a Pilot Proof-of-Concept Study

Circulation ◽  
2021 ◽  
Vol 144 (Suppl_2) ◽  
Author(s):  
Alexandra Weissman ◽  
Mariam Bramah Lawani ◽  
Thomas Rohan ◽  
Clifton W CALLAWAY
Keyword(s):  
Patient Care ◽  
Pathogen Detection ◽  
Bacterial Pathogen ◽  
Improve Patient Care ◽  
Antibiotic Administration ◽  
Nanopore Sequencing ◽  
Proof Of Concept ◽  
Chi Square ◽  
Significant Difference ◽  
Improve Patient

Introduction: Pneumonia is common after OHCA but is difficult to diagnose in the first 72 hours following ROSC, this results in early untargeted antibiotic administration based on non-specific imaging and laboratory findings. Antibiotic resistance is rising, is influenced by untargeted antibiotic administration, and can increase patient morbidity and mortality as well as healthcare costs. Precision methods of bacterial pathogen detection in OHCA patients are needed to improve patient care. This proof-of-concept pilot study aimed to assess feasibility of bacterial pathogen sequencing and comparability of sequencing results to clinical culture after OHCA. Methods: Blood and bronchoalveolar lavage (BAL) were obtained from residual clinical specimens collected within 12 hours of ROSC. Bacterial DNA was extracted using the Qiagen PowerLyzer PowerSoil DNA kit, sequenced using the MinION nanopore sequencer, and analyzed with Oxford Nanopore Technologies’ EPI2ME bioinformatics software. Sequencing results were compared to culture results using McNemar’s chi-square statistic. Study-defined pneumonia was based on presence of at least two characteristics within 72 hours of ROSC: fever (temperature ≥38°C); persistent leukocytosis >15,000 or leukopenia <3,500 for 48 hours; persistent chest radiography infiltrates for 48 hours per clinical radiology read; bacterial pathogen cultured. Results: We enrolled 38 consecutive OHCA subjects: mean age 61.8 years (18.0); 16 (42%) female; 25 (66%) White, 7 (18%) Black, 6 (16%) “Other” race; 7 subjects (18%) survived and 31 (82%) died; 16 (42%) subjects had pneumonia. Sequencing results were available in 12 hours while culture results were available in 48-72 hours after collection. There was a non-significant difference in the proportion of the same pathogens identified for each method per McNemar’s chi-square: p = 0.38, difference of 0.095 (-0.095, 0.286). Conclusions: Nanopore sequencing detects pathogenic bacteria comparable to clinical microbiologic culture and in less time. This technology can produce a paradigm shift in early bacterial pathogen detection in OHCA survivors, which can improve patient care. The technology is applicable to other patient populations and for viral and fungal pathogens.

scholarly journals Bacteriophage functional genomics and its role in bacterial pathogen detection

2013 ◽  
Vol 12 (4) ◽  
pp. 354-365 ◽  
Author(s):  
J. Klumpp ◽  
D. E. Fouts ◽  
S. Sozhamannan
Keyword(s):  
Functional Genomics ◽  
Pathogen Detection ◽  
Bacterial Pathogen

A fully integrated bacterial pathogen detection system based on count-on-a-cartridge platform for rapid, ultrasensitive, highly accurate and culture-free assay

2020 ◽  
Vol 152 ◽  
pp. 112007 ◽  
Author(s):  
Won-Il Lee ◽  
Younghyeon Park ◽  
Sajal Shrivastava ◽  
Taekeon Jung ◽  
Montri Meeseepong ◽  
...  
Keyword(s):  
Pathogen Detection ◽  
Detection System ◽  
Bacterial Pathogen ◽  
Fully Integrated

scholarly journals Cultivation-independent approach for the direct detection of bacteria in human clinical specimens as a tool for analysing culture-negative samples: a prospective study

SpringerPlus ◽  
2016 ◽  
Vol 5 (1) ◽  
Author(s):  
Ma. Guadalupe Aguilera-Arreola ◽  
Marcos Daniel Martínez-Peña ◽  
Fabiola Hernández-Martínez ◽  
Sara R. Juárez Enriques ◽  
Beatriz Rico Verdín ◽  
...  
Keyword(s):  
Prospective Study ◽  
Direct Detection ◽  
Clinical Specimens ◽  
A Prospective Study ◽  
Culture Negative

scholarly journals Depletion of Human DNA in Spiked Clinical Specimens for Improvement of Sensitivity of Pathogen Detection by Next-Generation Sequencing

2016 ◽  
Vol 54 (4) ◽  
pp. 919-927 ◽  
Author(s):  
Mohammad R. Hasan ◽  
Arun Rawat ◽  
Patrick Tang ◽  
Puthen V. Jithesh ◽  
Eva Thomas ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Pathogen Detection ◽  
Clinical Samples ◽  
Metagenomic Sequencing ◽  
Next Generation ◽  
Simple Method ◽  
Clinical Specimens ◽  
Human Dna ◽  
Pathogen Dna ◽  
Generation Sequencing

Next-generation sequencing (NGS) technology has shown promise for the detection of human pathogens from clinical samples. However, one of the major obstacles to the use of NGS in diagnostic microbiology is the low ratio of pathogen DNA to human DNA in most clinical specimens. In this study, we aimed to develop a specimen-processing protocol to remove human DNA and enrich specimens for bacterial and viral DNA for shotgun metagenomic sequencing. Cerebrospinal fluid (CSF) and nasopharyngeal aspirate (NPA) specimens, spiked with control bacterial and viral pathogens, were processed using either a commercially available kit (MolYsis) or various detergents followed by DNase prior to the extraction of DNA. Relative quantities of human DNA and pathogen DNA were determined by real-time PCR. The MolYsis kit did not improve the pathogen-to-human DNA ratio, but significant reductions (>95%;P< 0.001) in human DNA with minimal effect on pathogen DNA were achieved in samples that were treated with 0.025% saponin, a nonionic surfactant. Specimen preprocessing significantly decreased NGS reads mapped to the human genome (P< 0.05) and improved the sensitivity of pathogen detection (P< 0.01), with a 20- to 650-fold increase in the ratio of microbial reads to human reads. Preprocessing also permitted the detection of pathogens that were undetectable in the unprocessed samples. Our results demonstrate a simple method for the reduction of background human DNA for metagenomic detection for a broad range of pathogens in clinical samples.

Comparison of conventional culture with SeptiFast real-time PCR for microbial pathogen detection in clinical specimens other than blood

2011 ◽  
Vol 60 (12) ◽  
pp. 1774-1778 ◽  
Author(s):  
Antonella Mencacci ◽  
Christian Leli ◽  
Angela Cardaccia ◽  
Paolo Montagna ◽  
Amedeo Moretti ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Pathogen Detection ◽  
Clinical Specimens ◽  
Microbial Pathogen ◽  
Conventional Culture

In-House Validation of a Rinse–Membrane Filtration Method for Processing Fresh Produce Samples for Downstream Cultural Detection of Salmonella, Escherichia coli O157:H7, and Listeria

2020 ◽  
Vol 83 (9) ◽  
pp. 1592-1597
Author(s):  
LAURA E. TIJERINA-RODRÍGUEZ ◽  
LUISA SOLÍS-SOTO ◽  
NORMA HEREDIA ◽  
JUAN S. LEÓN ◽  
LEE-ANN JAYKUS ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Membrane Filtration ◽  
Pathogen Detection ◽  
Fresh Produce ◽  
Bacterial Pathogen ◽  
Relative Sensitivity ◽  
Good Alternative ◽  
Detection Methods ◽  
Filtration Method ◽  
Relative Specificity

ABSTRACT More efficient sampling and detection methods of pathogens on fresh produce are needed. The purpose of this study was to compare a novel rinse–membrane filtration method (RMFM) to a more traditional sponge rubbing or stomaching method in processing jalapeño peppers and cantaloupe samples for detection of Escherichia coli, Salmonella enterica, and Listeria monocytogenes. For jalapeño peppers inoculated with 106, 104, and 102 CFU of each pathogen and cantaloupes inoculated at 106 and 104 CFU, all pathogens were detected in all (100%) samples by RMFM at a 10-mL filtration volume, as well as by the stomacher and sponge rubbing methods. However, for cantaloupe inoculated at 102 CFU, detection differed by pathogen: S. enterica (20% RMFM, 60% stomacher, and 20% sponge), L. monocytogenes (40% RMFM, 60% stomacher, and 20% sponge), and E. coli O157:H7 (100% RMFM, 75% stomacher, and 75% sponge). When RMFM was compared with the other methods, in accordance with guidelines in the International Organization for Standardization 16140:2003 protocol, it produced values &gt;95% in relative accuracy, relative specificity, and relative sensitivity. Overall, the RMFM performed similar to or better than the homogenization and sponge surface rubbing methods and is a good alternative for processing large numbers of produce samples for bacterial pathogen detection.

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