scholarly journals Rapid ultrasensitive diagnosis of pneumonia caused by Acinetobacter baumannii using a combination of enrichment and phage-based qPCR assay

2020 ◽  
Author(s):  
Jun Luo ◽  
Mengwei Jiang ◽  
Jin Xiong ◽  
Junhua Li ◽  
Hongping Wei ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Enrichment Culture ◽  
Detection System ◽  
Sputum Sample ◽  
High Sensitivity ◽  
Rapid Identification ◽  
Clinical Method ◽  
Real Time Quantitative Pcr ◽  
Live Bacteria ◽  
Infection Patient

Abstract Background Accurate and rapid identification of ventilator associated or nosocomial pneumonia caused by Acinetobacter baumannii ( A. baumannii ) could improve the treatment.Methods In current study, we developed a phage-based real-time quantitative PCR (qPCR) combined with enrichment culture for rapid and specific detection of viable A. baumannii in sputum from lung infections. Through short-term plate incubation, bacteria can be enriched and the DNA polymerase reaction disturbance of sputum can be decreased greatly. This approach is based on detecting phage replication in live A. baumannii cells through Taqman qPCR.Results Through the built detection system, down to 1 CFU of A. baumannii can be detected within 6 h in spiked sputum samples without any steps of bacteria isolation and DNA extraction. The established method was then applied to detecting both A. baumannii in simulated sputum with 100% agreement with the spiked amount of the bacteria and one clinical sputum sample from an 80-year-old male lung infection patient caused by A. baumannii with perfect accuracy, demonstrating that the assay developed in this study has the merits of high rapidity, high sensitivity, good specificity and being able to detect live bacteria not dead bacteria.Conclusions The assay is a potentially clinical method for diagnosis of bacterial pneumonia infection caused by A. baumannii or other bacterial infection in sputum or complicated samples through switching to other types of phages.

scholarly journals Multiplex and On-site PCR detection of swine diseases based on the microfluidic chip system

2020 ◽  
Author(s):  
Yan Jiang ◽  
Shan Jiang ◽  
Yue Wu ◽  
Bin Zhou ◽  
Kaiming Wang ◽  
...  
Keyword(s):  
High Throughput ◽  
Microfluidic Chip ◽  
Detection System ◽  
Sampling Site ◽  
High Sensitivity ◽  
Pcr Detection ◽  
Clinical Samples ◽  
Pathogenic Microorganisms ◽  
Real Time Quantitative Pcr ◽  
Inspection And Quarantine

Abstract Background： At present, the process of inspection and quarantine starts with sampling at the customs port, continues with transporting the samples to the central laboratory for inspection experiments, and ends with the inspected results being fed back to the port. This process had the risks of degradation of biological samples and generation of pathogenic microorganisms and did not meet the rapid on-site detection demand because it took a rather long time. Therefore, it is urgently needed to develop a rapid and high-throughput detection assay of pathogenic microorganisms at the customs port. The aim of this study was to develop a microfluidic chip to rapidly detect swine pathogenic microorganisms with high-throughput and higher accuracy. Moreover, this chip will decrease the risk of spreading infection during transportation.Results: A series of experiments were performed to establish a microfluidic chip. The resulting data showed that the positive nucleic acid of four swine viruses were detected by using a portable and rapid microfluidic PCR system, which could achieve a on-site real-time quantitative PCR detection. Furthermore, the detection results of eight clinical samples were obtained within an hour. The detection limit of this microfluidic PCR detection system was as low as 1 copies/μL. The results showed that the high sensitivity and specificity of this chip system in disease detection played an important role in customs inspection and quarantine during customs clearance.Conclusion: The microfluidic PCR detection system established in this study could meet the requirement for rapid detection of samples at the customs port. This chip could avoid the risky process of transporting the samples from the sampling site to the testing lab, and drastically reduce the inspection cycle. Moreover, it would enable parallel inspections on one chip, which greatly raised the efficiency of inspection.

scholarly journals Multiplex and On-site PCR detection of swine diseases based on the microfluidic chip system

2021 ◽  
Author(s):  
Yan Jiang ◽  
Shan Jiang ◽  
Yue Wu ◽  
Bin Zhou ◽  
Kaiming Wang ◽  
...  
Keyword(s):  
High Throughput ◽  
Microfluidic Chip ◽  
Detection System ◽  
Sampling Site ◽  
High Sensitivity ◽  
Pcr Detection ◽  
Clinical Samples ◽  
Pathogenic Microorganisms ◽  
Real Time Quantitative Pcr ◽  
Inspection And Quarantine

Abstract Background: At present, the process of inspection and quarantine starts with sampling at the customs port, continues with transporting the samples to the central laboratory for inspection experiments, and ends with the inspected results being fed back to the port. This process had the risks of degradation of biological samples and generation of pathogenic microorganisms and did not meet the rapid on-site detection demand because it took a rather long time. Therefore, it is urgently needed to develop a rapid and high-throughput detection assay of pathogenic microorganisms at the customs port. The aim of this study was to develop a microfluidic chip to rapidly detect swine pathogenic microorganisms with high-throughput and higher accuracy. Moreover, this chip will decrease the risk of spreading infection during transportation.Results: A series of experiments were performed to establish a microfluidic chip. The resulting data showed that the positive nucleic acid of four swine viruses were detected by using a portable and rapid microfluidic PCR system, which could achieve a on-site real-time quantitative PCR detection. Furthermore, the detection results of eight clinical samples were obtained within an hour. The detection limit of this microfluidic PCR detection system was as low as 1 copies/μL. The results showed that the high sensitivity and specificity of this chip system in disease detection played an important role in customs inspection and quarantine during customs clearance.Conclusion: The microfluidic PCR detection system established in this study could meet the requirement for rapid detection of samples at the customs port. This chip could avoid the risky process of transporting the samples from the sampling site to the testing lab, and drastically reduce the inspection cycle. Moreover, it would enable parallel inspections on one chip, which greatly raised the efficiency of inspection.

scholarly journals Multiplex and On-site PCR detection of swine diseases based on the microfluidic chip system

2020 ◽  
Author(s):  
Yan Jiang ◽  
Shan Jiang ◽  
Yue Wu ◽  
Bin Zhou ◽  
Kaiming Wang ◽  
...  
Keyword(s):  
High Throughput ◽  
Microfluidic Chip ◽  
Detection System ◽  
Sampling Site ◽  
High Sensitivity ◽  
Pcr Detection ◽  
Clinical Samples ◽  
Pathogenic Microorganisms ◽  
Real Time Quantitative Pcr ◽  
Inspection And Quarantine

Abstract Background： At present, the process of inspection and quarantine starts with sampling at the customs port, continues with transporting the samples to the central laboratory for inspection experiments, and ends with the inspected results being fed back to the port. This process had the risks of degradation of biological samples and generation of pathogenic microorganisms and did not meet the rapid on-site detection demand because it took a rather long time. Therefore, it is urgently needed to develop a rapid and high-throughput detection assay of pathogenic microorganisms at the customs port. The aim of this study was to develop a microfluidic chip to rapidly detect swine pathogenic microorganisms with high-throughput and higher accuracy. Moreover, this chip will decrease the risk of spreading infection during transportation.Results: A series of experiments were performed to establish a microfluidic chip. The resulting data showed that the positive nucleic acid of four swine viruses were detected by using a portable and rapid microfluidic PCR system, which could achieve a on-site real-time quantitative PCR detection. Furthermore, the detection results of eight clinical samples were obtained within an hour. The detection limit of this microfluidic PCR detection system was as low as 1 copies/μL. The results showed that the high sensitivity and specificity of this chip system in disease detection played an important role in customs inspection and quarantine during customs clearance.Conclusion: The microfluidic PCR detection system established in this study could meet the requirement for rapid detection of samples at the customs port. This chip could avoid the risky process of transporting the samples from the sampling site to the testing lab, and drastically reduce the inspection cycle. Moreover, it would enable parallel inspections on one chip, which greatly raised the efficiency of inspection.

scholarly journals Taqman real-time quantitative PCR for identification of western flower thrip ( Frankliniella occidentalis ) for plant quarantine

2010 ◽  
Vol 6 (4) ◽  
pp. 555-557 ◽  
Author(s):  
K. S. Huang ◽  
S. E. Lee ◽  
Y. Yeh ◽  
G. S. Shen ◽  
E. Mei ◽  
...  
Keyword(s):  
Real Time ◽  
Quantitative Pcr ◽  
Ribosomal Rna ◽  
Frankliniella Occidentalis ◽  
Detection System ◽  
High Sensitivity ◽  
Plant Viruses ◽  
Agricultural Products ◽  
Morphological Identification ◽  
Real Time Quantitative Pcr

Western flower thrip ( Frankliniella occidentalis ) is a major global pest of agricultural products. It directly damages crops through feeding, oviposition activity or transmission of several plant viruses. We describe a Taqman real-time quantitative PCR detection system, which can rapidly identify F. occidentalis from thrips larvae to complement the traditional morphological identification. The data showed that our detection system targeted on the ribosomal RNA gene regions of F. occidentalis has high sensitivity and specificity. The rapid method can be used for on-site testing of samples at ports-of-entry in the future.

scholarly journals Multiplex and On-site PCR detection of swine diseases based on the microfluidic chip system

2020 ◽  
Author(s):  
Yan Jiang ◽  
Shan Jiang ◽  
Yue Wu ◽  
Bin Zhou ◽  
Kaiming Wang ◽  
...  
Keyword(s):  
High Throughput ◽  
Microfluidic Chip ◽  
Detection System ◽  
Sampling Site ◽  
High Sensitivity ◽  
Pcr Detection ◽  
Clinical Samples ◽  
Pathogenic Microorganisms ◽  
Real Time Quantitative Pcr ◽  
Inspection And Quarantine

Abstract Background： At present, the process of inspection and quarantine starts with sampling at the customs port, continues with transporting the samples to the central laboratory for inspection experiments, and ends with the inspected results being fed back to the port. This process had the risks of degradation of biological samples and generation of pathogenic microorganisms and did not meet the rapid on-site detection demand because it took a rather long time. Therefore, it is urgently needed to develop a rapid and high-throughput detection assay of pathogenic microorganisms at the customs port. The aim of this study was to develop a microfluidic chip to rapidly detect swine pathogenic microorganisms with high-throughput and higher accuracy. Moreover, this chip will decrease the risk of spreading infection during transportation. Results: A series of experiments were performed to establish a microfluidic chip. The resulting data showed that the positive nucleic acid of four swine viruses were detected by using a portable and rapid microfluidic PCR system, which could achieve a on-site real-time quantitative PCR detection. Furthermore, the detection results of eight clinical samples were obtained within an hour. The detection limit of this microfluidic PCR detection system was as low as 1 copies/μL. The results showed that the high sensitivity and specificity of this chip system in disease detection played an important role in customs inspection and quarantine during customs clearance. Conclusion: The microfluidic PCR detection system established in this study could meet the requirement for rapid detection of samples at the customs port. This chip could avoid the risky process of transporting the samples from the sampling site to the testing lab, and drastically reduce the inspection cycle. Moreover, it would enable parallel inspections on one chip, which greatly raised the efficiency of inspection.

A Multiplex PCR Assay Mediated by Universal Primers for the Detection of Adulterated Meat in Mutton

2019 ◽  
Vol 82 (2) ◽  
pp. 325-330 ◽  
Author(s):  
WANWAN LIU ◽  
XIAONAN WANG ◽  
JING TAO ◽  
BANGSHENG XI ◽  
MAN XUE ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Detection System ◽  
Meat Products ◽  
Pcr Primers ◽  
Rapid Identification ◽  
Food Samples ◽  
Detection Sensitivity ◽  
Universal Primers ◽  
Multiplex Pcr Assay ◽  
Multiple Pcr

ABSTRACT This study aimed to establish a multiplex PCR detection system mediated by “universal primers,” which would be able to determine whether mutton meat contained nonmutton ingredients from rats, foxes, and ducks. Based on the sequence variation of specific mitochondrial genes, nine different multiplex PCR primers were designed, and four kinds of meat products were rapidly identified by electrophoresis using an optimized multiplex PCR system based on the molecular weight differences of the amplified products. Multiplex PCR applications optimized for meat food source from food samples for testing was used to verify the accuracy of the identification method. The results showed that the primers in multiple PCR system mediated by universal primers could be used for the rapid identification of rat, fox, duck, and sheep meat in mutton products, and the detection sensitivity could reach 0.05 ng/μL. The identification of food samples validated the practical value of this method. Therefore, a multiplex PCR system mediated by universal primers was established, which can be used to quickly identify the origin of animal ingredients from rats, foxes, and ducks in mutton products.

scholarly journals Lytic Myophage Abp53 Encodes Several Proteins Similar to Those Encoded by Host Acinetobacter baumannii and Phage phiKO2

2011 ◽  
Vol 77 (19) ◽  
pp. 6755-6762 ◽  
Author(s):  
Chia-Ni Lee ◽  
Tsai-Tien Tseng ◽  
Juey-Wen Lin ◽  
Yung-Chieh Fu ◽  
Shu-Fen Weng ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Burst Size ◽  
Sputum Sample ◽  
Polyacrylamide Gel Electrophoresis ◽  
Multiple Drug ◽  
Lytic Phage ◽  
Tail Fiber ◽  
Drug Resistant ◽  
Content Type ◽  
Multiple Drug Resistant

ABSTRACTAcinetobacter baumanniiis an important Gram-negative opportunistic pathogen causing nosocomial infections. The emergence of multiple-drug-resistantA. baumanniiisolates has increased in recent years. Directed toward phage therapy, a lytic phage ofA. baumannii, designated Abp53, was isolated from a sputum sample in this study. Abp53 has an isometric head and a contractile tail with tail fibers (belonging toMyoviridae), a latent period of about 10 min, and a burst size of approximately 150 PFU per infected cell. Abp53 could completely lyse 27% of theA. baumanniiisolates tested, which were all multiple drug resistant, but not other bacteria. Mg2+enhanced the adsorption and productivity of, and host lysis by, Abp53. Twenty Abp53 virion proteins were visualized in SDS-polyacrylamide gel electrophoresis, with a 47-kDa protein being the predicted major capsid protein. Abp53 has a double-stranded DNA genome of 95 kb. Sequence analyses of a 10-kb region revealed 8 open reading frames. Five of the encoded proteins, including 3 tail components and 2 hypothetical proteins, were similar to proteins encoded byA. baumanniistrain ACICU. ORF1176 (one of the tail components, 1,176 amino acids [aa]), which is also similar to tail protein gp21 ofKlebsiellaphage phiKO2, contained repeated domains similar to those within the ACICU_02717 protein ofA. baumanniiACICU and gp21. These findings suggest a common ancestry and horizontal gene transfer during evolution. As phages can expand the host range by domain duplication in tail fiber proteins, repeated domains in ORF1176 might have a similar significance in Abp53.

Abstract 4336: Interrogating the Human Myocardial Interstitium during Myocardial Arrest and Reperfusion: Dynamic Changes in Cytokines and Protease Activity

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Robert E Stroud ◽  
Christine N Koval ◽  
Isabelle Gengler ◽  
Anne M Deschamps ◽  
John S Ikonomidis ◽  
...  
Keyword(s):  
Detection System ◽  
Ischemia Reperfusion ◽  
Dynamic Assessment ◽  
High Sensitivity ◽  
Fluorescent Detection ◽  
Suspension Array ◽  
Cytokine Analysis ◽  
And Function ◽  
Inflammatory Molecules ◽  
Pulmonary Bypass

Background. Cytokines, such as the interleukins (IL1β, IL2, IL6) and tumor necrosis factor (TNF) can modulate myocardial structure and function with ischemia/reperfusion (I/R) but dynamic assessment of these biological molecules within the human myocardial interstitium with I/R has not been performed, and the inter-relationship to matrix metalloproteinases activity (MMPact) remains unexplored. Accordingly, a fluorogenic microdialysis method was used to simultaneously measure myocardial interstitial cytokine levels and MMPact in patients during and following I/R. Methods . MMPact was measured in patients (n=13) undergoing cardio-pulmonary bypass (CPB) at baseline, during myocardial arrest and CPB (on-CPB), and immediately following reperfusion and separation from CPB (post-CPB) by a validated in-line microdialysis fluorescent detection system. Myocardial interstitial fluid was subjected to cytokine analysis by high sensitivity multiplex suspension array. Results . Interstitial MMPact increased by over 30% post-CPB and was accompanied by a specific change in cytokine profiles (Figure ). The classical pro-inflammatory molecules such as TNF and IL6 were either not detectable or unchanged, whereas IL1β and IL2 which can be proinflammatory, were increased. Conclusions. These unique results demonstrated that a dynamic cytokine signature occurs within the human myocardial interstitium following I/R and is temporally related to heightened MMP activity. Direct interrogation of the human myocardial interstitium may provide a unique insight into critical signaling pathways which may evoke adverse structural and functional events following I/R.

Compact Detection System for High Sensitivity Hydrogen Profiling of Materials by Nuclear Reaction Analysis

2009 ◽  
Author(s):  
Daniel Keith Marble ◽  
Ben Urban ◽  
Jose Pacheco ◽  
Floyd D. McDaniel ◽  
Barney L. Doyle
Keyword(s):  
Nuclear Reaction ◽  
Detection System ◽  
High Sensitivity ◽  
Nuclear Reaction Analysis
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