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

Oubre, Cherie; Birmele, Michele N.; Castro, Victoria; Venkateswaran, Kasthuri; Vaishampayan, Parag; Jones, K. U.; Singhal, Adesh; Johnston, Albert S.; Roman, Monsi C.; Ozbolt, Tamra A; Jett, Daniel X; Roberts, Michael S.; Ott, C. Mark

doi:10.2514/6.2013-3314

Cited by 8 publications

(10 citation statements)

References 1 publication

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“…Among the target-based quantification approaches, the quantitative PCR (qPCR) represents a popular technique for the in-flight water quality monitoring (Oubre et al, 2013). The costs (Ibekwe et al, 2002;LaGier et al, 2004).…”

Section: Target-based Techniques For Detection and Quantification Of mentioning

confidence: 99%

Water and Microbial Monitoring Technologies towards the Near Future Space Exploration

Amalfitano¹,

Levantesi²,

Copetti³

et al. 2019

Preprint

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Space exploration is demanding longer lasting human missions and water resupply from Earth will become increasingly unrealistic. In a near future, the spacecraft water monitoring systems will require technological advances to promptly identify and counteract contingent events of waterborne microbial contamination, posing health risks to astronauts with lowered immune responsiveness. The search for bio-analytical approaches, alternative to those applied on Earth by cultivation-dependent methods, is pushed by the compelling need to limit waste disposal and avoid microbial regrowth from analytical carryovers. Prospective technologies will be selected only if first validated in a flight-like environment, by following basic principles, advantages, and limitations beyond their current applications on Earth. Starting from the water monitoring activities applied on the International Space Station, we provide a critical overview of the nucleic acid amplification-based approaches (i.e., loop-mediated isothermal amplification, quantitative PCR, and high-throughput sequencing) and early-warning methods for total microbial load assessments (i.e., ATP-metry, flow cytometry), already used at a high readiness level aboard crewed space vehicles. Our findings suggest that the forthcoming space applications of mature technologies will be necessarily bounded by a compromise between analytical performances (e.g., speed to results, identification depth, reproducibility, multiparametricity) and detrimental technical requirements (e.g., reagent usage, waste production, operator skills, crew time). As space exploration progresses toward extended missions to Moon and Mars, miniaturized systems that also minimize crew involvement in their end-to-end operation are likely applicable on the long-term and suitable for the in-flight water and microbiological research.

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Section: Target-based Techniques For Detection and Quantification Of mentioning

confidence: 99%

Water and Microbial Monitoring Technologies towards the Near Future Space Exploration

Amalfitano¹,

Levantesi²,

Copetti³

et al. 2019

Preprint

View full text Add to dashboard Cite

show abstract

“…In 2016, astronauts on board the ISS successfully deployed the Wetlab-2 research platform to quantitatively analyze gene expression via RT-qPCR. However, although dedicated equipment has been developed to facilitate the extraction and purification of nucleic acids, sample preparation in microgravity remains a problem, mainly due to the risk of sample carryover and contamination by the ISS environment (Oubre et al, 2013).…”

Section: Implantable Biosensors Glucowizzard™mentioning

confidence: 99%

Advanced biosensors for monitoring astronauts’ health during long-duration space missions

Roda

Mirasoli

Guardigli

et al. 2018

Biosensors and Bioelectronics

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“…The extracted nucleic acids captured on magnetic particles can be eluted into an elution buffer (e.g., Tris-HCl or water) or directly into master mix. In space, nucleic acids can be eluted into a Cepheid SmartCycler (already installed inside the ISS) PCR reaction tube to perform real-time PCR [ 8 , 27 , 28 ]. For space applications, the lyophilization of PCR and isothermal amplification reagents means that the preloaded reaction mix can be stored for months without refrigeration.…”

Section: Introductionmentioning

confidence: 99%

The Development of a 3D Printer-Inspired, Microgravity-Compatible Sample Preparation Device for Future Use Inside the International Space Station

et al. 2023

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Biological testing on the International Space Station (ISS) is necessary in order to monitor the microbial burden and identify risks to crew health. With support from a NASA Phase I Small Business Innovative Research contract, we have developed a compact prototype of a microgravity-compatible, automated versatile sample preparation platform (VSPP). The VSPP was built by modifying entry-level 3D printers that cost USD 200–USD 800. In addition, 3D printing was also used to prototype microgravity-compatible reagent wells and cartridges. The VSPP’s primary function would enable NASA to rapidly identify microorganisms that could affect crew safety. It has the potential to process samples from various sample matrices (swab, potable water, blood, urine, etc.), thus yielding high-quality nucleic acids for downstream molecular detection and identification in a closed-cartridge system. When fully developed and validated in microgravity environments, this highly automated system will allow labor-intensive and time-consuming processes to be carried out via a turnkey, closed system using prefilled cartridges and magnetic particle-based chemistries. This manuscript demonstrates that the VSPP can extract high-quality nucleic acids from urine (Zika viral RNA) and whole blood (human RNase P gene) in a ground-level laboratory setting using nucleic acid-binding magnetic particles. The viral RNA detection data showed that the VSPP can process contrived urine samples at clinically relevant levels (as low as 50 PFU/extraction). The extraction of human DNA from eight replicate samples showed that the DNA extraction yield is highly consistent (there was a standard deviation of 0.4 threshold cycle when the extracted and purified DNA was tested via real-time polymerase chain reaction). Additionally, the VSPP underwent 2.1 s drop tower microgravity tests to determine if its components are compatible for use in microgravity. Our findings will aid future research in adapting extraction well geometry for 1 g and low g working environments operated by the VSPP. Future microgravity testing of the VSPP in the parabolic flights and in the ISS is planned.

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

Cited by 8 publications

References 1 publication

Water and Microbial Monitoring Technologies towards the Near Future Space Exploration

Water and Microbial Monitoring Technologies towards the Near Future Space Exploration

Advanced biosensors for monitoring astronauts’ health during long-duration space missions

The Development of a 3D Printer-Inspired, Microgravity-Compatible Sample Preparation Device for Future Use Inside the International Space Station

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