Engineering topics

Castro, V. A.

Publications and source records attributed to Castro, V. A..

10 records

NASA NTRS2014

The Impact of Apollo-Era Microbiology on Human Space Flight

The microbiota of crewmembers and the spacecraft environment contributes significant risk to crew health during space flight missions. NASA reduces microbial risk with various mitigation methods that originated during the Apollo Program and continued to evolve through subsequent programs: Skylab, Shuttle, and International Space Station (ISS). A quarantine of the crew and lunar surface samples, within the Lunar Receiving Laboratory following return from the Moon, was used to prevent contamination with unknown extraterrestrial organisms. The quarantine durations for the crew and lunar samples were 21 days and 50 days, respectively. A series of infections among Apollo crewmembers resulted in a quarantine before launch to limit exposure to infectious organisms. This Health Stabilization Program isolated the crew for 21 days before flight and was effective in reducing crew illness. After the program developed water recovery hardware for Apollo spacecraft, the 1967 National Academy of Science Space Science Board recommended the monitoring of potable water. NASA implemented acceptability limits of 10 colony forming units (CFU) per mL and the absence of viable E. coli, anaerobes, yeasts, and molds in three separate 150 mL aliquots. Microbiological investigations of the crew and spacecraft environment were conducted during the Apollo program, including the Apollo-Soyuz Test Project and Skylab. Subsequent space programs implemented microbial screening of the crew for pathogens and acceptability limits on spacecraft surfaces and air. Microbiology risk mitigation methods have evolved since the Apollo program. NASA cancelled the quarantine of the crew after return from the lunar surface, reduced the duration of the Health Stabilization Program; and implemented acceptability limits for spacecraft surfaces and air. While microbial risks were not a main focus of the early Mercury and Gemini programs, the extended duration of Apollo flights resulted in the increased scrutiny of impact of the space flight environment on crew health. The lessons learned during that era of space flight continue to impact microbiology risk mitigation in space programs today.

Elliott, T. F↗

NASA NTRS2012

Microbial Monitoring Challenges and Needs for Mars Applications

The monitoring of microorganisms will be an important part of a mission to Mars. Microbial monitoring equipment will be needed to look for the presence of microorganisms on the planet, to confirm that planetary protection measures are working, to monitor the health of plants, bioreactors and humans living in a habitat and to monitor the performance of the life support systems that will keep them alive during their stay on Mars. Coordinating the different microbial monitoring needs during the early days of mission planning, can provide NASA with equipment that could meet more than one need while also providing complementary analysis options, which can enhance the research capabilities. The early coordination between the different NASA groups that will need microbial monitoring equipment on the surface of Mars, could also make the mission more affordable, as development of the needed equipment could be potentially cost shared.

Roman, M. C.↗

NASA NTRS2012

Surface, Water, and Air Biocharacterization (SWAB) Flight Experiment

The determination of risk from infectious disease during spaceflight missions is composed of several factors including both the concentration and characteristics of the microorganisms to which the crew are exposed. Thus, having a good understanding of the microbial ecology aboard spacecraft provides the necessary information to mitigate health risks to the crew. While preventive measures are taken to minimize the presence of pathogens on spacecraft, medically significant organisms have been isolated from both the Mir and International Space Station (ISS). Historically, the method for isolation and identification of microorganisms from spacecraft environmental samples depended upon their growth on culture media. Unfortunately, only a fraction of the organisms may grow on a specific culture medium, potentially omitting those microorganisms whose nutritional and physical requirements for growth are not met. To address this bias in our understanding of the ISS environment, the Surface, Water, and Air Biocharacterization (SWAB) Flight Experiment was designed to investigate and develop monitoring technology to provide better microbial characterization. For the SWAB flight experiment, we hypothesized that environmental analysis using non-culture-based technologies would reveal microorganisms, allergens, and microbial toxins not previously reported in spacecraft, allowing for a more complete health assessment. Key findings during this experiment included: a) Generally, advanced molecular techniques were able to reveal a few organisms not recovered using culture-based methods; however, there is no indication that current monitoring is "missing" any medically significant bacteria or fungi. b) Molecular techniques have tremendous potential for microbial monitoring, however, sample preparation and data analysis present challenges for spaceflight hardware. c) Analytical results indicate that some molecular techniques, such as denaturing gradient gel electrophoresis (DGGE), can be much less sensitive than culture-based methods. d) More sensitive molecular techniques, such as quantitative polymerase chain reaction (QPCR), were able to identify viral DNA from ISS environments, suggesting potential transfer of the organism between crewmembers. In addition, the hardware selected for this experiment represented advances for next-generation sample collection. The advanced nature of this collection hardware was noted, when the Sartorius MD8 Air Port air sampler from the SWAB experiment remained on board ISS at the request of JAXA investigators, who intend to use it in completion of their microbial ecology experiment.

Castro, V. A.↗

NASA NTRS2012

Next Generation Microbiology Requirements

As humans continue to explore deep into space, microorganisms will travel with them. The primary means to mitigate the risk of infectious disease are a combination of prudent spacecraft design and rigorous operational controls. The effectiveness of these methods are evaluated by microbiological monitoring of spacecraft, food, water, and the crew that is performed preflight, in-flight, and post-flight. Current NASA requirements associated with microbiological monitoring are based on culture-based methodology where microorganisms are grown on a semi-solid growth medium and enumerated. Subsequent identification of the organisms requires specialized labor and large equipment, which historically has been performed on Earth. Requirements that rely strictly on culture-based units limit the use of non-culture based monitoring technology. Specifically, the culture-based "measurement criteria" are Colony Forming Units (CFU, representing the growth of one microorganism at a single location on the agar medium) per a given volume, area, or sample size. As the CFU unit by definition is culture-based, these requirements limit alternative technologies for spaceflight applications. As spaceflight missions such as those to Mars extend further into space, culture-based technology will become difficult to implement due to the (a) limited shelf life of the culture media, (b) mass/volume necessary to carry these consumables, and (c) problems associated with the production of biohazardous material in the habitable volume of the spacecraft. In addition, an extensive amount of new knowledge has been obtained during the Space Shuttle, NASA-Mir, and International Space Station Programs, which gave direction for new or modified microbial control requirements for vehicle design and mission operations. The goal of this task is to develop and recommend a new set of requirements for vehicle design and mission operations, including microbiological monitoring, based upon "lessons learned" and new technology. During 2011, this study focused on evaluating potable water requirements by assembling a forum of internal and external experts from NASA, other federal agencies, and academia. Key findings from this forum included: (1) Preventive design and operational strategies should be stringent and the primary focus of NASA's mitigation efforts, as they are cost effective and can be attained with conventional technology. (2) Microbial monitoring hardware should be simple and must be able to measure the viability of microorganisms in a sample. Multiple monitoring technologies can be utilized as long as at the microorganisms being identified can also be confirmed as viable. (3) Evidence showing alterations in the crew immune function and microbial virulence complicates risk assessments and creates the need for very conservative requirements. (4) One key source of infectious agents will always be the crew, and appropriate preventative measures should be taken preflight. (5) Water systems should be thoroughly disinfected (sterilized if possible) preflight and retain a residual biocide throughout the mission. Future forums will cover requirements for other types of samples, specifically spaceflight food and environmental samples, such as vehicle air and vehicle and cargo surfaces. An interim report on the potable water forum has been delivered to the Human Research Program with a final report on the recommendations for all sample types being delivered in September 2013.

Ott, C. M.↗

NASA NTRS2011

Latent Virus Reactivation in Space Shuttle Astronauts

Latent virus reactivation was measured in 17 astronauts (16 male and 1 female) before, during, and after short-duration Space Shuttle missions. Blood, urine, and saliva samples were collected 2-4 months before launch, 10 days before launch (L-10), 2-3 hours after landing (R+0), 3 days after landing (R+14), and 120 days after landing (R+120). Epstein-Barr virus (EBV) DNA was measured in these samples by quantitative polymerase chain reaction. Varicella-zoster virus (VZV) DNA was measured in the 381 saliva samples and cytomegalovirus (CMV) DNA in the 66 urine samples collected from these subjects. Fourteen astronauts shed EBV DNA in 21% of their saliva samples before, during, and after flight, and 7 astronauts shed VZV in 7.4% of their samples during and after flight. It was interesting that shedding of both EBV and VZV increased during the flight phase relative to before or after flight. In the case of CMV, 32% of urine samples from 8 subjects contained DNA of this virus. In normal healthy control subjects, EBV shedding was found in 3% and VZV and CMV were found in less than 1% of the samples. The circadian rhythm of salivary cortisol measured before, during, and after space flight did not show any significant difference between flight phases. These data show that increased reactivation of latent herpes viruses may be associated with decreased immune system function, which has been reported in earlier studies as well as in these same subjects (data not reported here).

Mehta, S. K.↗

NASA NTRS2007

A Comprehensive Characterization of Microorganisms and Allergens in Spacecraft Environment

The determination of risk from infectious disease during long-duration missions is composed of several factors including (1) the host#s susceptibility, (2) the host#s exposure to the infectious disease agent, and (3) the concentration of the infectious agent, and (4) the characteristics of the infectious agent. While stringent steps are taken to minimize the transfer of potential pathogens to spacecraft, several medically significant organisms have been isolated from both the Mir and International Space Station (ISS). Historically, the method for isolation and identification of microorganisms from spacecraft environmental samples depended upon their growth on culture media. Unfortunately, only a fraction of the organisms may grow on a culture medium, potentially omitting those microorganisms whose nutritional and physical requirements for growth are not met. Thus, several pathogens may not have been detected, such as Legionella pneumophila, the etiological agent of Legionnaire#s disease. We hypothesize that environmental analysis using non-culture-based technologies will reveal microorganisms, allergens, and microbial toxins not previously reported in spacecraft, allowing for a more complete health assessment. The development of techniques for this flight experiment, operationally named SWAB, has already provided advances in NASA laboratory processes and beneficial information toward human health risk assessment. The first accomplishment of the SWAB experiment was the incorporation of 16S ribosomal DNA sequencing for the identification of bacteria. The use of this molecular technique has increased bacterial speciation of environmental isolates from previous flights three fold compared to conventional methodology. This increased efficiency in bacterial speciation provides a better understanding of the microbial ecology and the potential risk to the crew. Additional SWAB studies focused on the use of molecular-based DNA fingerprinting using repetitive sequencebased polymerase chain reaction (rep-PCR). This technology has allowed contamination tracking of microorganisms between crewmembers and their environment. This study not only demonstrated that ISS has a greater diversity of organisms than originally expected, but also provided insight into possible routes of infection to the crew. Additional ground-based studies used rep-PCR and protein based assays to determine the potential of methicillin resistant Staphylococcus aureus (MRSA) aboard ISS. MRSA has become increasingly common on Earth and pose a treatment problem for infections during flight. While no MRSA have been isolated from ISS to date, the mecA gene product that is responsible for methicillin resistance was isolated in other Staphylococcus species aboard ISS suggesting a potential of MRSA through gene transfer. Using improved sample collection technologies, flight sampling for SWAB was initiated in August 2006 and should continue through spring of 2007. The focus of these flight samples is the collection of DNA for evaluation by Denaturing Gradient Gel Electrophoresis (DGGE). Unlike other techniques, DGGE does not depend on any microbial growth on culture media allowing a more comprehensive assessment of the spacecraft interior. This study should provide insight into the true microbial ecology that is experienced by the crew during flight. This information will lead toward an accurate microbial risk assessment to help set flight requirements to protect the safety, health, and performance of the crew.

Ott, C. M.↗

NASA NTRS2006

Spacecraft Environment May Reduce Resistance To Infection

Living and working in a spacecraft exposes the crew to a unique environment. This environment includes microgravity, increased radiation, chemical and biological contamination, and a variety of stressors. Disturbances in this balance are often manifested by diminished immunity in astronauts/cosmonauts. Reactivation of Epstein- Barr virus (EBV), cytomegalovirus (CMV), and varicella-zoster virus (VZV) has been used as an indicator of immune status. Reactivation of EBV and VZV were detected and quantified in saliva. CMV was measured in urine. The DNA was extracted using a Qiagen Inc. kit and viral DNA was detected by real time polymerase chain reaction (PCR) based assay with Taqman 7700 (PE Biosystems). Patterns of Epstein-Barr virus (EBV) reactivation in 32 astronauts and 18 healthy age-matched control subjects were characterized by quantifying EBV shedding. Saliva samples were collected before, during, and after 10 space shuttle missions of 5 to 14 d duration. Of 1398 saliva specimens from 32 astronauts, 314 (23%) were positive for EBV DNA. Examination by flight phase showed that 29% of the saliva specimens collected from 28 astronauts before flight were positive for EBV DNA, as were 16% of those collected from 25 astronauts during flight and 16% of those collected after flight from 23 astronauts. The mean number of EBV copies/mL from samples taken during the flights was 417, ten-fold greater (p < 0.05) than the copies from the preflight (40) and post flight (44) phases. In contrast, the control subjects shed EBV DNA with a frequency of 3.7% and mean EBV copies of 40 per mL of saliva. Ten days before flight and on landing day, titers of antibody to EBV viral capsid antigen were significantly (p < 0.05) greater than baseline levels. Increases in the number of viral copies and in the amount of EBV-specific antibody were consistent with EBV reactivation before, during, and after space flight. Similarly, CMV and VZV reactivation increased in response to space flight conditions. Data indicates that space flight is a unique stress environment that may produce stress-induced changes in the host-microbe relationship resulting in increased risk of infection.

Pierson, Duane L.↗

NASA NTRS2005

The Evaluation of Methicillin Resistance in Staphylococcus aboard the International Space Station

The International Space Station (ISS) represents a semi-closed environment with a high level of crewmember interaction. As community-acquired methicillin-resistant Staphylococcus aureus (MRSA) has emerged as a health concern in environments with susceptible hosts in close proximity, an evaluation of isolates of clinical and environmental Staphylococcus aureus and coagulase negative Staphylococcus was performed to determine if this trend was also present in astronauts aboard ISS or the space station itself. Rep-PCR fingerprinting analysis of archived ISS isolates confirmed our earlier studies indicating a transfer of S. aureus between crewmembers. In addition, this fingerprinting also indicated a transfer between crewmembers and their environment. While a variety of S. aureus were identified from both the crewmembers and the environment, phenotypic evaluations indicated minimal methicillin resistance. However, positive results for the Penicillin Binding Protein, indicative of the presence of the mecA gene, were detected in multiple isolates of archived Staphylococcus epidermidis and Staphylococcus haemolyticus. Phenotypic analysis of these isolates confirmed their resistance to methicillin. While MRSA has not been isolated aboard ISS, the potential exists for the transfer of the gene, mecA, from coagulase negative environmental Staphylococcus to S. aureus creating MRSA strains. This study suggests the need to expand environmental monitoring aboard long duration exploration spacecraft to include antibiotic resistance profiling.

Ott, C. M.↗

NASA NTRS2004

Microbial Characterization During the Early Habitation of the International Space Station

An evaluation of the microbiota from air, water, and surface samples provided a baseline of microbial characterization onboard the International Space Station (ISS) to gain insight into bacterial and fungal contamination during the initial stages of construction and habitation. Using 16S genetic sequencing and rep-PCR, 63 bacterial strains were isolated for identification and fingerprinted for microbial tracking. Of the bacterial strains that were isolated and fingerprinted, 19 displayed similarity to each other. The use of these molecular tools allowed for the identification of bacteria not previously identified using automated biochemical analysis and provided a clear indication of the source of several ISS contaminants. Strains of Bradyrhizobium and Sphingomonas unable to be identified using sequencing were identified by comparison of rep-PCR DNA fingerprints. Distinct DNA fingerprints for several strains of Methylobacterium provided a clear indication of the source of an ISS water supply contaminant. Fungal and bacterial data acquired during monitoring do not suggest there is a current microbial hazard to the spacecraft, nor does any trend indicate a potential health risk. Previous spacecraft environmental analysis indicated that microbial contamination will increase with time and will require continued surveillance. Copyright 2004 Springer-Verlag.

Fungi/genetics↗

NASA NTRS2004

A Survey of Staphylococcus sp and its Methicillin Resistance aboard the International Space Station

Background: Within the past few years, methicillin-resistant Staphylococcus aureus has emerged in environments with susceptible hosts in close proximity, such as hospitals and nursing homes. As the International Space Station (ISS) represents a semi-closed environment with a high level of crewmember interaction, an evaluation of isolates of clinical and environmental Staphylococcus aureus and coagulase negative Staphylococcus was performed to determine if this trend was also present in astronauts occupying ISS or on surfaces of the space station itself. Methods: Identification of isolates was completed using VITEK (GPI cards, BioMerieux), 16S ribosomal DNA analysis (MicroSeq 500, ABI), and Rep-PCR DNA fingerprinting (Divemilab, Bacterial Barcodes). Susceptibility tests were performed using VITEK (GPS-105 cards, BioMerieux) and resistance characteristics were evaluated by testing for the presence of the mecA gene (PBP2' MRSA test kit, Oxoid). Results: Rep-PCR analysis indicated the transfer of S. aureus between crewmembers and between crewmembers and ISS surfaces. While a variety of S. aureus were identified from both the crewmembers and environment, evaluations of the microbial population indicated minimal methicillin resistance. Results of this study indicated that within the semi-closed ISS environment, transfer of bacteria between crewmembers and their environment has been occurring, although there was no indication of a high concentration of methicillin resistant Staphylococcus species. Conclusions: While this study suggests that the spread of methicillin resistant S. aureus is not currently a concern aboard ISS, the increasing incidence of Earth-based antibiotic resistance indicates a need for continued clinical and environmental monitoring.

Bassinger, V. J.↗