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Detection of Microorganisms Onboard the International Space Station Using an Electronic Nose

Ulrich Reidt
Ulrich Reidt
, 
Andreas Helwig
Andreas Helwig
, 
Gerhard Müller
Gerhard Müller
, 
Lutz Plobner
Lutz Plobner
, 
Veronika Lugmayr
Veronika Lugmayr
, 
Sergey Kharin
Sergey Kharin
, 
Yuri Smirnov
Yuri Smirnov
, 
Natalia Novikova
Natalia Novikova
, 
Joachim Lenic
Joachim Lenic
, 
Viktor Fetter
Viktor Fetter
 and 
Thomas Hummel
Thomas Hummel
   | Jul 21, 2020
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Article Category: Research Article
Published Online: Jul 21, 2020
Page range: 89 - 111
DOI: https://doi.org/10.2478/gsr-2017-0013
Keywords
© 2017 Ulrich Reidt et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Figure 1

(A) Cultivation station with 12 custom-built gas vessels in which the organisms are grown under controlled gaseous environments. The E-Nose is connected on the left for measurement of the cultures. (B) Gas vessels with the culture of Bacillus subtilis in liquid tryptic soy broth media.
(A) Cultivation station with 12 custom-built gas vessels in which the organisms are grown under controlled gaseous environments. The E-Nose is connected on the left for measurement of the cultures. (B) Gas vessels with the culture of Bacillus subtilis in liquid tryptic soy broth media.

Figure 2

Illustration of the E-Nose and the Target Book that were used for experiments at the ISS. (A) The E-Nose is connected with an air sampler via a transfer line. Attached to the air sampler is a charcoal filter. Figure (B) The Target Book with four different materials: A1–A3, non-anodized aluminum; B1–B3, Nomex; C1–C3, printed circuit board material; and D1–D3, cable labeling material.
Illustration of the E-Nose and the Target Book that were used for experiments at the ISS. (A) The E-Nose is connected with an air sampler via a transfer line. Attached to the air sampler is a charcoal filter. Figure (B) The Target Book with four different materials: A1–A3, non-anodized aluminum; B1–B3, Nomex; C1–C3, printed circuit board material; and D1–D3, cable labeling material.

Figure 3

Training of cosmonaut Roman Romanenko in the mock-up of the Zvezda Service Module at Star City, Russia.
Training of cosmonaut Roman Romanenko in the mock-up of the Zvezda Service Module at Star City, Russia.

Figure 4

Surface Pipette Kit used for swabbing the surfaces of the three locations of measurements. Cotton swabs (Q-tip style) from left to right: Q1: working table, Q2: sleeping cabin, and Q3: toilet lid.
Surface Pipette Kit used for swabbing the surfaces of the three locations of measurements. Cotton swabs (Q-tip style) from left to right: Q1: working table, Q2: sleeping cabin, and Q3: toilet lid.

Figure 5

(A) Photograph of the International Space Station (ISS). The Russian segment with the Zvezda Service Module in which the measurements were performed is labeled with a red arrow (photograph from: http://www.nasa.gov). (B) Illustration of the three measurement locations at the Zvezda Service Module: a) working table, b) sleeping cabin, and c) toilet lid. (C) Measurement of working table with the E-Nose. The astronaut presses the air sampler against the working table. (D) Negative control measurements in space. The cosmonaut measures the sterile Target Book.
(A) Photograph of the International Space Station (ISS). The Russian segment with the Zvezda Service Module in which the measurements were performed is labeled with a red arrow (photograph from: http://www.nasa.gov). (B) Illustration of the three measurement locations at the Zvezda Service Module: a) working table, b) sleeping cabin, and c) toilet lid. (C) Measurement of working table with the E-Nose. The astronaut presses the air sampler against the working table. (D) Negative control measurements in space. The cosmonaut measures the sterile Target Book.

Figure 6

(A) Comparison of the measurements from the ISS (measurement locations and Target Book in pink rhombs) with the four teaching organisms Bacillus subtilis (green squares), Staphylococcus warneri (red rhombs), Penicillium expansum (blue triangle), and Aspergillus versicolor (turquoise triangles). In score plot PC1/2, the space measurements were located close to the cluster of B. subtilis and the negative controls (gray spheres). (6B) Representation of the measurements at ISS (a–c) in score plot PC1/3 related to all four teaching organisms (B. subtilis, S. warneri, P. expansum, and A. versicolor). The E-Nose measurements at the ISS were located distal from all teaching species. Only the measurement at position c (toilet seat) in test series 2 (2-c) is located outside of the remaining seven E-Nose measurements (a–c). (C) In score plot PC2/3, results from space measurements were presented versus the teaching organisms. Measurements performed at the ISS build a cluster separate from all other measurements. (D) Detailed description of the E-Nose measurements obtained at Zvezda Service Module illustrated in score plot PC3/4. From left to right; 1-b: campaign 1, location b (sleeping cabin); 3-a: campaign 3, location a (working table); 2-a: campaign 2, location a (working table); 1-a: campaign 1, location a (working table); 2-b: campaign 2, location b (sleeping cabin); 3-c: campaign 3, location c (toilet lid); 1-c: campaign 1, location c (toilet lid); 2-c: campaign 2, location c (toilet lid).
(A) Comparison of the measurements from the ISS (measurement locations and Target Book in pink rhombs) with the four teaching organisms Bacillus subtilis (green squares), Staphylococcus warneri (red rhombs), Penicillium expansum (blue triangle), and Aspergillus versicolor (turquoise triangles). In score plot PC1/2, the space measurements were located close to the cluster of B. subtilis and the negative controls (gray spheres). (6B) Representation of the measurements at ISS (a–c) in score plot PC1/3 related to all four teaching organisms (B. subtilis, S. warneri, P. expansum, and A. versicolor). The E-Nose measurements at the ISS were located distal from all teaching species. Only the measurement at position c (toilet seat) in test series 2 (2-c) is located outside of the remaining seven E-Nose measurements (a–c). (C) In score plot PC2/3, results from space measurements were presented versus the teaching organisms. Measurements performed at the ISS build a cluster separate from all other measurements. (D) Detailed description of the E-Nose measurements obtained at Zvezda Service Module illustrated in score plot PC3/4. From left to right; 1-b: campaign 1, location b (sleeping cabin); 3-a: campaign 3, location a (working table); 2-a: campaign 2, location a (working table); 1-a: campaign 1, location a (working table); 2-b: campaign 2, location b (sleeping cabin); 3-c: campaign 3, location c (toilet lid); 1-c: campaign 1, location c (toilet lid); 2-c: campaign 2, location c (toilet lid).

Figure 7

(A) Additional teaching of the E-Nose by Pseudomonas geniculate and comparison with the measurements from ISS (a–c). In score plot PC1/2 P. geniculate (green rhombs) builds a distinct cluster at the opposite quadrant of the space measurements. (B) Comparison of measurements in space and the odor determined from Saccharomyces cerevisiae (orange squares) illustrated in the score plot PC1/2. Odor signals spread out in the right direction. (C) Teaching of the E-Nose by the yeast strain Rhodotorula mucilaginosa versus measurements performed at the ISS (a–c). Both signals coincide in one common cluster in the score plot PC1/2. (D) Score plot PC3/4 of the signals from R. mucilaginosa and the measurements in space. The signals coincide.
(A) Additional teaching of the E-Nose by Pseudomonas geniculate and comparison with the measurements from ISS (a–c). In score plot PC1/2 P. geniculate (green rhombs) builds a distinct cluster at the opposite quadrant of the space measurements. (B) Comparison of measurements in space and the odor determined from Saccharomyces cerevisiae (orange squares) illustrated in the score plot PC1/2. Odor signals spread out in the right direction. (C) Teaching of the E-Nose by the yeast strain Rhodotorula mucilaginosa versus measurements performed at the ISS (a–c). Both signals coincide in one common cluster in the score plot PC1/2. (D) Score plot PC3/4 of the signals from R. mucilaginosa and the measurements in space. The signals coincide.

Figure 8

(A) Electrophoretic analysis of PCR fragments generated with the primer pair NS1/NS2 from DNA of the surface samples Q1 (working table), Lanes 1–3; Q2 (sleeping cabin), Lanes 4–6; and Q3 (toilet lid), Lanes 7–9. As a negative control, 0.85% (w/v) NaCl solution in place of a sample was processed to show that there is no unspecific yeast DNA background in any of the reagents used (Lanes 10–12). As no template control (NTC), 5 μL of pure water was added to the PCR assay (Lane 13). The size marker was applied in Lane 14 and labeled on the left-hand side. (B) Real-time PCR specifically analyzing the DNA from Rhodotorula mucilaginosa that was isolated from the three surface samples Q1 (working table), Q2 (sleeping cabin), and Q3 (toilet lid). For sample Q2, a strong signal could be detected. No presence of R. mucilaginosa DNA was seen in Q1 and Q3. As a negative control, the whole method was performed with pure water instead of a sample to assure that there is no unspecific R. mucilaginosa DNA in any of the reagents used. For the positive control, DNA from a pure culture of R. mucilaginosa was isolated and added to the qPCR assay. To check whether there is any unspecific reaction with other Rhodotorula species, DNA from a pure culture of R. glutinis was used. For the NTC, 5 μL of pure water were analyzed with the qPCR assay.
(A) Electrophoretic analysis of PCR fragments generated with the primer pair NS1/NS2 from DNA of the surface samples Q1 (working table), Lanes 1–3; Q2 (sleeping cabin), Lanes 4–6; and Q3 (toilet lid), Lanes 7–9. As a negative control, 0.85% (w/v) NaCl solution in place of a sample was processed to show that there is no unspecific yeast DNA background in any of the reagents used (Lanes 10–12). As no template control (NTC), 5 μL of pure water was added to the PCR assay (Lane 13). The size marker was applied in Lane 14 and labeled on the left-hand side. (B) Real-time PCR specifically analyzing the DNA from Rhodotorula mucilaginosa that was isolated from the three surface samples Q1 (working table), Q2 (sleeping cabin), and Q3 (toilet lid). For sample Q2, a strong signal could be detected. No presence of R. mucilaginosa DNA was seen in Q1 and Q3. As a negative control, the whole method was performed with pure water instead of a sample to assure that there is no unspecific R. mucilaginosa DNA in any of the reagents used. For the positive control, DNA from a pure culture of R. mucilaginosa was isolated and added to the qPCR assay. To check whether there is any unspecific reaction with other Rhodotorula species, DNA from a pure culture of R. glutinis was used. For the NTC, 5 μL of pure water were analyzed with the qPCR assay.

Determination of colony forming units (CFU) from the washout of swabs.

Culture mediaQ1 (working table), CFU/mLQ2 (sleeping cabin), CFU/mLQ3 (toilet lid), CFU/mL
MacConkey Agar000
Mannitol Salt Agar000
R2A Agar1403.9E+040
Sabouraud 4% Glucose Agar14.0E+040
Tryptic Soy Agar21.9E+073.7E+04
Yeast Extract Agar202.5E+042.8E+04
∑: CFU/mL1.63E+021.91E+076.50E+04

Theoretical prediction of the MVOCs from Saccharomyces cerevisiae compared to sensor compound sensitivity of the E-Nose.

Saccharomyces cerevisiaeE-Nose
MVOCChem. classificationNo. of sensors and compounds sensitivity
1-PropanolAlcoholSensor 1: Aromatic compound, Alkane
2-Ethyl-1-hexanolAlcoholSensor 2: Nitrogen oxides, Alcohols, Aromatic compounds
2-Methyl-1-propanolAlcoholSensor 3: Aromatic compound, Alkane
2-PropanolAlcoholSensor 4: Hydrogen
EthanolAlcoholSensor 5: Aromatic compounds, Alkane
Phenethyl AlcoholAlcoholSensor 6: Methane, Alcohol
2-(Ethenyloxy)-ethanolAlcohol, Ether, AlkeneSensor 7: Inorganic sulfur compounds, Terpenes, Sulfur-containing organic compounds, Aromatic compound
1,2-Benzenedicarboxylic AcidAcidSensor 8: Alcohol, Aromatic compound
2-Methyl Propanoic AcidAcidSensor 9: Aromatic compounds and inorganic sulfur compounds as H2S
2-MethylbutanoateAcidSensor 10: Methane and aliphatic organic compounds
3-Methylbutanoic AcidAcid
Acetic AcidAcid
2-ButanoneKetone
2-PentanoneKetone
2-PropanoneKetone
1,9-Dimethylspiro(4,5)decaneAlkane
UndecaneAlkane
Alpha-limoneneAlkene, Terpene
3-Methyl butanalAldehyde
AcetaldehydeAldehyde
Acetic Acid Ethenyl EsterEster
Ethyl AcetateEster
PyrazinePyrazine
2,5-DimethylpyrazinePyrazine
Dimethyl DisulfideSulfide
Ortho-DimethylbenzeneBenzenoid

Genetic identification of microorganisms in 25 pure cultures from selected colonies by BLAST analysis.

Q1 (working table)Q2 (sleeping cabin)Q3 (toilet lid)
Penicillium expansumfAgrobacterium tumefaciensb,−Bacillus subtilisb,+
Staphylococcus epidermidisb,+Bacillus agaradhaerensb,+Staphylococcus warnerib,+
Streptococcus salivariusb,+Bacillus atrophaeusb,+Staphylococcus epidermidisb,+
Fusarium oxysporumf
Micrococcus luteusb,+
Pseudomonas geniculatab,−
Staphylococcus hominisb,+
Stenotrophomonas maltophiliab,−
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