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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 91 records · Page 5
All sourcesNASA NTRSDOE OSTI
DOE OSTI2018

Multi-site evaluation of the LN34 pan-lyssavirus real-time RT-PCR assay for post-mortem rabies diagnostics

Rabies is a fatal zoonotic disease that requires fast, accurate diagnosis to prevent disease in an exposed individual. The current gold standard for post-mortem diagnosis of human and animal rabies is the direct fluorescent antibody (DFA) test. While the DFA test has proven sensitive and reliable, it requires high quality antibody conjugates, a skilled technician, a fluorescence microscope and diagnostic specimen of sufficient quality. The LN34 pan-lyssavirus real-time RT-PCR assay represents a strong candidate for rabies post-mortem diagnostics due to its ability to detect RNA across the diverse Lyssavirus genus, its high sensitivity, its potential for use with deteriorated tissues, and its simple, easy to implement design. Here, we present data from a multi-site evaluation of the LN34 assay in 14 laboratories. A total of 2,978 samples (1,049 DFA positive) from Africa, the Americas, Asia, Europe, and the Middle East were tested. The LN34 assay exhibited low variability in repeatability and reproducibility studies and was capable of detecting viral RNA in fresh, frozen, archived, deteriorated and formalin-fixed brain tissue. The LN34 assay displayed high diagnostic specificity (99.68%) and sensitivity (99.90%) when compared to the DFA test, and no DFA positive samples were negative by the LN34 assay. The LN34 assay produced definitive findings for 80 samples that were inconclusive or untestable by DFA; 29 were positive. Five samples were inconclusive by the LN34 assay, and only one sample was inconclusive by both tests. Furthermore, use of the LN34 assay led to the identification of one false negative and 11 false positive DFA results. Together, these results demonstrate the reliability and robustness of the LN34 assay and support a role for the LN34 assay in improving rabies diagnostics and surveillance.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

Influenza A viral burst size from thousands of infected single cells using droplet quantitative PCR (dqPCR)

An important aspect of how viruses spread and infect is the viral burst size, or the number of new viruses produced by each infected cell. Surprisingly, this value remains poorly characterized for influenza A virus (IAV), commonly known as the flu. In this study, we screened tens of thousands of cells using a microfluidic method called droplet quantitative PCR (dqPCR). The high-throughput capability of dqPCR enabled the measurement of a large population of infected cells producing progeny virus. By measuring the fully assembled and successfully released viruses from these infected cells, we discover that the viral burst sizes for both the seasonal H3N2 and the 2009 pandemic H1N1 strains vary significantly, with H3N2 ranging from 10 1 to 10 4 viruses per cell, and H1N1 ranging from 10 1 to 10 3 viruses per cell. Some infected cells produce average numbers of new viruses, while others generate extensive number of viruses. In fact, we find that only 10% of the single-cell infections are responsible for creating a significant portion of all the viruses. This small fraction produced approximately 60% of new viruses for H3N2 and 40% for H1N1. On average, each infected cell of the H3N2 flu strain produced 709 new viruses, whereas for H1N1, each infected cell produced 358 viruses. This novel method reveals insights into the flu virus and can lead to improved strategies for managing and preventing the spread of viruses.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2022

End-User Perspectives on Using Quantitative Real-Time PCR and Genomic Sequencing in the Field

Quantitative real-time PCR and genomic sequencing have become mainstays for performing molecular detection of biological threat agents in the field. There are notional assessments of the benefits, disadvantages, and challenges that each of these technologies offers according to findings in the literature. However, direct comparison between these two technologies in the context of field-forward operations is lacking. Most market surveys, whether published in print form or provided online, are directed to product manufacturers who can address their respective specifications and operations. One method for comparing these technologies is surveying end-users who are best suited for discussing operational capabilities, as they have hands-on experience with state-of-the-art molecular detection platforms and protocols. These end-users include operators in military defense and first response, as well as various research scientists in the public sector such as government and service laboratories, private sector, and civil society such as academia and nonprofit organizations performing method development and executing these protocols in the field. Our objective was to initiate a survey specific to end-users and their feedback. We developed a questionnaire that asked respondents to (1) determine what technologies they currently use, (2) identify the settings where the technologies are used, whether lab-based or field-forward, and (3) rate the technologies according to a set list of criteria. Of particular interest are assessments of sensitivity, specificity, reproducibility, scalability, portability, and discovery power. This article summarizes the findings from the end-user perspective, highlighting technical and operational challenges.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS1991

STS-37 The Payload bay door closing at PCR Pad B

The primary objective of the STS-37 mission was to deploy the Gamma Ray Observatory. The mission was launched at 9:22:44 am on April 5, 1991, onboard the space shuttle Atlantis. This videotape shows the payload bay doors being closed. Included are views of the Gamma Ray Observatory in the payload bay, and the clean room operations in the Payload Changeout Room (PCR).

Source record↗
NASA NTRS1991

STS-37 Payload Gamma Ray Observatory Pad-B in PCR

The primary objective of the STS-37 mission was to deploy the Gamma Ray Observatory. The mission was launched at 9:22:44 am on April 5, 1991, onboard the space shuttle Atlantis. This videotape shows the Gamma Ray Observatory being placed in the payload bay of the shuttle. The Payload Changeout Room (PCR) and the clean room operations required to place the payload in the bay are shown.

Source record↗
NASA NTRS2002

Parallel characterization of anaerobic toluene- and ethylbenzene-degrading microbial consortia by PCR-denaturing gradient gel electrophoresis, RNA-DNA membrane hybridization, and DNA microarray technology

A mesophilic toluene-degrading consortium (TDC) and an ethylbenzene-degrading consortium (EDC) were established under sulfate-reducing conditions. These consortia were first characterized by denaturing gradient gel electrophoresis (DGGE) fingerprinting of PCR-amplified 16S rRNA gene fragments, followed by sequencing. The sequences of the major bands (T-1 and E-2) belonging to TDC and EDC, respectively, were affiliated with the family Desulfobacteriaceae. Another major band from EDC (E-1) was related to an uncultured non-sulfate-reducing soil bacterium. Oligonucleotide probes specific for the 16S rRNAs of target organisms corresponding to T-1, E-1, and E-2 were designed, and hybridization conditions were optimized for two analytical formats, membrane and DNA microarray hybridization. Both formats were used to characterize the TDC and EDC, and the results of both were consistent with DGGE analysis. In order to assess the utility of the microarray format for analysis of environmental samples, oil-contaminated sediments from the coast of Kuwait were analyzed. The DNA microarray successfully detected bacterial nucleic acids from these samples, but probes targeting specific groups of sulfate-reducing bacteria did not give positive signals. The results of this study demonstrate the limitations and the potential utility of DNA microarrays for microbial community analysis.

Non-NASA Center↗
NASA NTRS1998

PCR cloning and characterization of multiple ADP-glucose pyrophosphorylase cDNAs from tomato

Four ADP-glucose pyrophosphorylase (AGP) cDNAs were cloned from tomato fruit and leaves by the PCR techniques. Three of them (agp S1, agp S2, and agp S3) encode the large subunit of AGP, the fourth one (agp B) encodes the small subunit. The deduced amino acid sequences of the cDNAs show very high identities (96-98%) to the corresponding potato AGP isoforms, although there are major differences in tissue expression profiles. All four tomato AGP transcripts were detected in fruit and leaves; the predominant ones in fruit are agp B and agp S1, whereas in leaves they are agp B and agp S3. Genomic southern analysis suggests that the four AGP transcripts are encoded by distinct genes.

Non-NASA Center↗
NASA NTRS1990

PCR amplification of 16S rDNA from lyophilized cell cultures facilitates studies in molecular systematics

The sequence of the major portion of a Bacillus cycloheptanicus strain SCH(T) 16S rRNA gene is reported. This sequence suggests that B. cycloheptanicus is genetically quite distinct from traditional Bacillus strains (e.g., B. subtilis) and may be properly regarded as belonging to a different genus. The sequence was determined from DNA that was produced by direct amplification of ribosomal DNA from a lyophilized cell pellet with straightforward polymerase chain reaction (PCR) procedures. By obviating the need to revive cell cultures from the lyophile pellet, this approach facilitates rapid 16S rDNA sequencing and thereby advances studies in molecular systematics.

Non-NASA Center↗
NASA NTRS2008

Mold Species in Dust from the International Space Station Identified and Quantified by Mold Specific Quantitative PCR

Dust was collected over a period of several weeks in 2007 from various HEPA filters in the U.S. Laboratory Module of the International Space Station (ISS). The dust was returned on the Space Shuttle Atlantis, mixed, sieved, and the DNA was extracted. Using a DNA-based method called mold specific quantitative PCR (MSQPCR), 39 molds were measured in the dust. Opportunistic pathogens Aspergillus flavus and A. niger and toxin producers Penicillium chrysogenum and P. brevicompactum were found at relatively high concentrations (compared to U.S. homes). No cells of the opportunistic pathogens A. fumigatus, A. terreus, Fusarium solani or Candida albicans were detected.

Vesper, Stephen J.↗
NASA NTRS2013

Microbial Monitoring of Common Opportunistic Pathogens by Comparing Multiple Real-time PCR Platforms for Potential Space Applications

The International Space Station is a closed environment with rotation of astronauts and equipment that each introduce their own microbial flora, it is necessary to monitor the air, surfaces, and water for microbial contamination. Current microbial monitoring includes labor- and time-intensive methods to enumerate total bacterial and fungal cells, with limited characterization, during in-flight testing. Although this culture-based method is sufficient for monitoring the International Space Station, on future long-duration missions more characterization will need to be performed during flight, as sample return and ground characterization may not be available.

Real time↗