Open came a o QR eade and
scan code o access his a icle
and o he esou ces online.
In Si u Real-Time Moni o ing o Asep ic D illing:
Lessons Lea ned om he A acama Ro e
As obiology D illing S udies Con amina ion Con ol
S a egy and Implemen a ion and Applica ion o he
Iceb eake Ma s Li e De ec ion Mission
Rosalba Bonacco si,
1,2
B ian Glass,
2
Me cedes Mo eno-Paz,
3
Mi iam Ga cı´a-Villadangos,
3
Kimbe ley Wa en-Rhodes,
1,2
Vic o Pa o,
3
Juan Manuel Manchado,
3
Ma y Be h Wilhelm,
2
and Ch is ophe P. McKay
2
Abs ac
In 2019, he A acama Ro e As obiology D illing S udies (ARADS) p ojec ield- es ed an au onomous o e -
moun ed obo ic d ill p o o ype o a 6-Sol li e de ec ion mission o Ma s (Iceb eake ). ARADS d illed Ma s-
like ma e ials in he A acama Dese (Chile), one o he mos li e-diminished egions on Ea h, whe e mi iga ing
con amina ion ans e in o li e-de ec ion ins umen s becomes c i ical. Ou Con amina ion Con ol S a egy
and Implemen a ion (CCSI) o he Sample Handling and T ans e Sys em (SHTS) ha dwa e (d ill, scoop and
unnels) included ou -o -simula ion p o ocol es ing (ou -o -sim) o ha dwa e decon amina ion and e i ica ion
du ing he 6-Sol simula ion (in-sim). The mos e ec i e i e-s ep decon amina ion combined sa e - o-use
s e ilan s (3%_hyd ogen-pe oxide-ac i a ed 5%_sodium-hypochlo i e), and in si u eal- ime e i ica ion by
adenosine iphospha e (ATP) and Signs o Li e De ec o (SOLID) Fluo escence Immunoassay cha ac e iza ion
o ha dwa e biobu den and ai bo ne con aminan s. The 20- o 40-min p o ocol enabled a 4-log biobu den
educ ion down o <0.1 moles ATP de ec ion limi ( unnels and d ill) o 0.2–0.7 moles (scoop) o o al ATP. The
(pos -cleaning) ha dwa e backg ound was 0.3 o 1–2 a omoles ATP/cm
2
(cleanliness benchma k backg ound
alues) equi alen o ca. 1–10 colony o ming uni (CFU)/cm
2
. Fu he , 60–100% o he in-sim ha dwa e back-
g ound was £3–4 bac e ial cells/cm
2
, he h eshold limi o Class <7 asep ic ope a ions. Ac oss he six Sols, he lux
o ai bo ne con aminan s o he d ill si es was *5and*22 amoles ATP/(cm
2
$day), accoun ing o an unex-
pec edly high Fluo escence In ensi y (FI) signal (FI: *6000) agains aqua ic cyanobac e ia, bu negligible an-
h opogenic con ibu ion. The SOLID immunoassay also de ec ed mic oo ganisms om mul iple habi a s ac oss he
A acama Dese (anoxic, alkaline/acidic mic oen i onmen s in hali e ields, playas, and allu ial ans) in bo h
ai bo ne and pos -cleaning ha dwa e backg ound. Finally, he ha dwa e ATP backg ound was 40–250 imes lowe
han he ATP in co es. Simila ly, he FI peaks (FI
max
) agains he mic obial axa and molecula bioma ke s de ec ed
in he pos -cleaned ha dwa e (FI: *1500–1600) we e 5–10 imes lowe han bioma ke s in d illed sedimen s,
excluding signi ican in e e ence wi h pu a i e bioma ke ound in co es. Simila p o ocols enable he acquisi ion
o con amina ion- ee ma e ials o ul a-sensi i e ins umen s analysis and he in eg i y o scien i ic esul s. Thei
applica ion can augmen ou scien i ic knowledge o he dis ibu ion o c yp ic li e on Ma s-like g ounds and
1
SETI Ins i u e, Moun ain View, Cali o nia, USA.
2
NASA Ames Resea ch Cen e , Mo e Field, Cali o nia, USA.
3
Depa men o Molecula E olu ion, Cen o de As obiologı
´a (INTA-CSIC), Mad id, Spain.
Rosalba Bonacco si e al., 2023; Published by Ma y Ann Liebe , Inc. This Open Access a icle is dis ibu ed unde he e ms o he
C ea i e Commons License (h p://c ea i ecommons.o g/licenses/by/4.0), which pe mi s un es ic ed use, dis ibu ion, and ep oduc ion in
any medium, p o ided he o iginal wo k is p ope ly c edi ed.
ASTROBIOLOGY
Volume 23, Numbe 12, 2023
Ma y Ann Liebe , Inc.
DOI: 10.1089/as .2022.0133
1303
suppo li e-de ec ion obo ic and human-ope a ed missions o Ma s. Key Wo ds: A acama—Mic obial educ ion—
Real- ime ATP assay—Robo ic d illing—Li e de ec ion—Ma s’Iceb eake Sample Handling and T ans e Sys em
(SHTS)—A acama Ro e As obiology D illing S udies (ARADS). As obiology 23, 1303–1336.
1. In oduc ion
Unlike Ea h, he ma ian su ace appea s li eless, wi h
he subsu ace p oposed as he bes a ge o de ec ing
li e. The A acama Ro e As obiology D illing S udies
(ARADS) sys em is a p o o ype o a u u e li e de ec ion
d ill mission o Ma s, namely he cu en ly planned Ice-
b eake Mission (Da e
´e al., 2013; McKay e al., 2013; Glass
e al., 2014; Mann, 2018). ARADS pe o med ield ials in
he hype -a id egion o he A acama Dese in he An o a-
gas a Region om 2016 o 2019 (Fig. 1), culmina ing in a
6-Sol end- o-end mission simula ion.
As obiology obo ic d illing missions on Ma s will ace
b oad issues bo h in o wa d plane a y p o ec ion (PP) om
e es ial mic obes su i ing s e iliza ion o he spacec a ’s
ha dwa e, which is he p ima y conce n (e.g., NAS, 2020;
NAS, 2019; NAS, 2018; NRC, 2006) and in he in eg i y o
sample analysis. The Commi ee on Space Resea ch (CO-
SPAR) PP policy o landed missions in a Ma ian Special
Region (Ca ego y IVc PP, p obabili y con amina ion: <10
-4
)
is he mos s ic equi ing ha dwa e biobu den con amina ion
con ol o o wa d con amina ion (COSPAR, 2020).
Spo es can emain long- e m iable in space (e.g., Ho -
neck e al., 2010, 2012; Hallswo h, 2021), be shed on he
su ace (Ma shall and Mancinelli, 2011), and en e he
unde g ound, becoming me abolically ac i e unde inc ea-
sed wa e ac i i y (Rummel e al., 2014; Smi h e al., 2017).
A seconda y conce n is mi iga ing he isk o e es ial
con aminan s ans e (spo es, amino acids) in o sample
analysis gene a ing alse posi i es—li e appea s o be de ec-
ed when he e is no —(Kminek e al., 2019), o p o ec ing
science da a in eg i y. Human-ope a ed con amina ion
emedia ion is a p ima y s ep o obo ic d illing mission
planning, beginning wi h clean oom ope a ions du ing
Spacec a Assembly, Tes , and Launch Ope a ions (ATLO).
The human in ol emen con inues du ing obo ic d illing
mission mock-ups, as he hands-o mission echnology eadi-
ness is no ye ma u e, equi ing ield enginee s o supe ise
FIG. 1. (A) ARADS 2019 S udy Si es. (B) Playa es si e (La . 2406¢08.3†S o Long. 7008¢17.5†W) abou 22 km om
he ield suppo headqua e , he Yungay S a ion. (C) Si e loca ion o ‘‘G een Pa o ’’ (GP) La . 245¢37.62†S; Long.
6954¢20.86†W). ARADS, A acama Ro e As obiology D illing S udies; GP, G een Pa o .
1304 BONACCORSI ET AL.
decon amina ion (Glass e al., 2023, As obiology his issue).
Thus, de eloping and ield- es ing human- ended cleaning e-
gimes o payload p o o ypes add essing PP knowledge gaps is
manda o y e en o a obo ic mission (e.g., NASA, 2011) and
can be ad anced by ield ials such as ARADS.
We de eloped ou Con amina ion Con ol S a egy and Im-
plemen a ion (CCSI) o e he 4-yea p ojec and in eg a ed
lessons lea ned om se e al as obiology and echnology ields
d illing simula ions, o example, he 2005 MARTE P ojec a
he Rio Tin o Pen
˜a de Hie o, Spain (Glass e al., 2006; Bo-
nacco si and S oke , 2008; Mille e al., 2008; Pa o e al., 2008;
S oke e al., 2008; Eigenb ode e al., 2009; Amado e al., 2014;
Cockell e al., 2018). These es s p o ided oppo uni ies o assess
and mi iga e long- and sho - e m biobu den buil -up a *6and
12 mon hs and e alua e cleaning p o ocols o e-con amina ed
deep-cleaned ha dwa e ha dwa e a e 1 day o use.
This pape desc ibes ield- easible and e ec i e p ac ices
o con amina ion isk educ ion du ing he ARADS mission
and consis s o se en sec ions.
Sec ion 1 in oduces ARADS sample d illing handling and
ans e ha dwa e. Sec ion 2 in o ms spacec a ha dwa e con-
amina ion con ol. Sec ion 3 ou lines an h opogenic, en i on-
men al, and c oss-con amina ion. Sec ion 4 desc ibes (1) he
gene al con amina ion emedia ion (du ing o -hou s o e ,
ins umen main enance, e c.); (2) mul i-s ep cleaning, mic o-
bial educ ion, and e i ica ion o he d ill, scoop, and unnels;
(3) mul i-s ep p o ocols implemen a ion o in-sim ope a ions;
and (4) sou ce- o-sink con amina ion o ai bo ne, sedimen a y,
and an h opogenic biomass o he d ill si es.
Sec ion 5 epo s he 2019 ARADS esul s. Sec ion 6
discusses alse nega i es in cleaning e i ica ion and en i-
onmen al and c oss-con amina ion issues unique o e es-
ial ials o a li e-de ec ion d illing mission o Ma s.
Sec ion 7 highligh s inno a i e elemen s s emming om
ARADS: (1) an e ec i e simpli ied decon amina ion p o-
ocol, (2) a sou ce- o-sink con amina ion app oach, (3) a e-
assessmen o con amina ion isks in he A acama Dese ,
(4) he no el use o he immunoassay-based li e de ec-
ion ool (Signs o Li e De ec o [SOLID]) o iden i ying
o wa d-con aminan s on Ma s, and (5) ha dwa e design o he
a m-moun ed scoop analog o he Phoenix soil acquisi ion
de ice o mi iga e c oss-con amina ion.
1.1. In oduc ion o ARADS
The ARADS p ojec was pa o NASA’s Plane a y Science
and Technology Th ough Analog Resea ch (PSTAR) P og am.
This p ojec explo es he science, ins umen a ion, and au oma-
ion in pe o ming a nea su ace ( o 1 m dep h) d ill ope a ion on
Ma s, using he hype -a id A acama (Cab ol e al., 2007; McKay
e al., 2003; Na a o-Gonzalez e al., 2003; Cab ol e al., 2007) as
an analog d ill si e (e.g., Glass e al., 2019, 2022; Mo eno-Paz
e al., 2019; S oke e al., 2023). The inal ARADS ield cam-
paign concluded in Sep embe 2019 wi h a 6-day simula ed
mission a a no el ‘‘landing si e’’ he ‘‘Playa’’ (Fig. 1), selec ed
o mul iple g ound- u h ield s udies (Glass e al., 2023, his
issue). The ARADS CCSI cons i u ed an in eg al pa o he o e
ield deploymen in bo h in-sim and ou -o -sim ope a ions.
The e a e ou majo subsys ems o he ARADS payload:
(1) The NASA Ames’ sel -na iga ing midsize o e
(KREX-2) as he mechanical in eg a ion pla o m o
he 1 m TRIDENT (The Regoli h and Ice D ill o
Explo a ion o New Te ains) d ill and he o he sub-
sys ems hos ed by he Ro e payload deck (Fig. 2A).
(2) The Honeybee Robo ics’ o a y-pe cussi e D ill,
whose s ing spi al coils can e ie e ca. 10–60 g o
geological cu ings om e e y 25 cm o d illed sub-
su ace (Fig. 4C).
(3) The Sample Handling and T ans e Sys em (SHTS) in-
cludes an a m-moun ed scoop analog o he Phoenix Icy
Soil Acquisi ion De ice (ISAD) shown in Fig. 2D (Bo-
ni z e al., 2008; Glass e al., 2023) and ou unnels on
he op o he o e payload deck, p o ec ing he science
payload unde nea h (Fig. 2A): h ee *12 cm diame e
ins umen - eeding unnels, and one 13 cm diame e one
(aka ‘‘Big_Fun’’) o deli e o e low samples in o a
s o age uni o pos - ield analysis (Fig. 2C).
(4) The science ins umen s included (i) The Signs o
Li e De ec o (SOLID) o p o o ype, buil by he
Spanish Cen o de As obiologia (CAB) (Pa o e al.,
2008) using luo esce mic oa ay immunoassay de-
ec ion o 450 biomolecula compounds (amino acids,
nucleic acids, p o eins, and polysaccha ides) a 1–2
pa s pe billion [ppb (ng/mL)] and whole cells, ha
is, 10
4
–10
5
cells/mL (Ri as e al., 2008); (ii) he
JPL’s Chemical Lap op- o me ly Mic o luidic Li e
Analyze (MILA) (Willis e al., 2015; Mo a e al.,
2020); (iii) a b assboa d e sion o he Phoenix’s We
Chemis y Labo a o y (WCL) o sal chemis y
analysis; and (i ) he NASA Godda d Space Fligh
Cen e ’s Linea Ion T ap Mass Spec ome e , o
LITMS o o ganic compounds de ec ion and iden i-
ica ion, which was ield es ed Ma ch 2019 (Cas illo
e al., 2023). Nex , we desc ibe he sample handling
and ans e o hese ins umen s.
1.2. Li e jou ney o a d illed sample
The ARADS’s ully au oma ed o e /d illing/sampling/
analysis sys em aimed a d i ing o loca ions o scien i ic
in e es and acqui ing geological ma e ial o geochemical
and biological s udy. The sample cap u e occu s in h ee
s eps. The s ainless-s eel d ill s ing, nes ed inside he sys-
em’s p ima y d ill, accesses he dep h o science in e es and
e ie es ca. 10–60 g o powde ed cu ings. Be o e he sed-
imen is b ough o he su ace, he a m-moun ed scoop
lowe s o he g ound o ecei e he d illed ma e ial (Fig. 2B).
Once su icien powde ed sedimen om he a ge dep h
en e s he scoop, his ises abo e he o e deck (Fig. 2A) o
deli e ma e ial in o he ins umen s’ unnels (Fig. 2C).
Each unnel d ops he cu ings in o 1 cm diame e holes
aligned wi h he en y po o he analy ical ools benea h he
o e payload deck (no isible). Finally, he powde le in
he scoop is d opped in o he ou h, la ge unnel (aka
‘‘Big_Fun’’) and ans e ed in o a p e-cleaned bo osilica e
glass ja as in p e ious ARADS campaigns (Wilhelm e al.,
2018). The unnel-ja uni la ches on o he o e payload
deck op (Fig. 2A).
The o e ins umen sui e immedia ely analyzes he
d opped samples. Al e na i ely, he ins umen is de ached,
b ough in o he labo a o y aile , and analyzed immedi-
a ely by he science eam. Only one ins umen eam sci-
en is can wo k a he dedica ed sani ized lab bench o
minimize con amina ion du ing sample analysis.
ARADS CONTAMINATION CONTROL STRATEGY IMPLEMENTATION 1305
Finally, samples in ja s a e s o ed in a clean box o la e
sub-sampling and analysis o he a chi ed co ed ma e ials
on eques by he science eam. Fo his pu pose, we ope -
a ed on he clean bench op using s e ilized s ainless-s eel
spa ulas in he dedica ed science aile (Fig. 9A). We
excluded a ew low-p io i y d illed in e als om he sim-
ula ion o main ain mission ideli y. Howe e , unlike on
Ma s, we did no disca d hese low-p io i y samples om
he d ill bu manually collec ed hem in labeled s e ile
Whi l-Pakbags o la e use by he science eam.
1.3. Rele ance o con amina ion con ol
Cleaning, mic obial educ ion, and e i ica ion we e un-
damen al o ARADS. Unlike he case o human and obo ic
explo a ion o Ma s Special Regions, he o wa d con ami-
na ion o he e es ial poly-ex eme A acama Dese poses
no eal conce n. Howe e , because he hype -a id su ace o
his egion is p edominan ly de oid o mic obes, ins u-
men s o de ec molecula indica o s o pu a i e li e mus
ope a e unde low de ec ion h esholds (high sensi i i y).
E en mino biobu den con amina ion du ing sample acqui-
si ion and ans e in o analysis could con amina e scien i ic
esul s and con use in e p e a ions. Biobu den is he numbe
o mic oo ganisms on he non-s e ile su ace ma e ial. S e -
iliza ion e e s o chemical o physical p ocedu es ha kill,
deac i a e, o emo e all li e- o ms (i.e., MHRA, 2006).
A e minally s e ilized su ace de ice is ‘‘s e ile’’ i he he-
o e ical p obabili y o a iable mic oo ganism being p esen is
equal o o less han 1 ·10
-6
(Lambe e al., 2021). NASA
p o ocols in ol e d y hea o mic obial educ ion (DHMR) and
chemical me hods when DHMR is no a ainable. Howe e ,
long- e m absolu e s e ili y is unachie able o is comp omised
a e a ew-minu e exposu e o a con amina ing en i onmen .
Fu he , ha dy mic oo ganisms can esis chemical ea -
men in clean ooms (Ho neck e al., 2012; Moissl-Eichinge
e al., 2015; Sp y e al., 2021) and e en use o ganic com-
pounds in cleaning eagen s as an ene gy sou ce (e.g., Mogul
e al., 2018). Field ials in ex eme en i onmen s lack clean-
oom acili ies, and con inuous human- ended ope a ions en-
hance he isk exposu e o biobu den con amina ion o he
ha dwa e payload and sample analysis pa hways.
FIG. 2. SHTS. (A) Payload deck’s
op hos ing he h ee sample deli e y
unnels and he la ge unnel (back-
g ound) o he ou low sample ans e
o he glass ja (no in place); (B) low-
e ed scoop ecei ing d illed sample;
(C) scoop abo e payload deck deli e -
ing cu ings o he unnel eeding he
SOLID ins umen below (no isible).
The glass ja is isible in he back-
g ound. (D) Phoenix ISAD design
simila o ARADS scoop (Pho o c edi :
NASA). ISAD, Icy Soil Acquisi ion
De ice; SHTS, Sample Handling
and T ans e Sys em; SOLID, Signs
o Li e De ec o .
1306 BONACCORSI ET AL.
The isk is enhanced unde a ime-cons ained schedule
o decon amina ion. Each Sol, he ARADS simula ion ini-
ially alloca ed only a 20-min ope a ional window o he
mic obial educ ion and e i ica ion cycle, making c i ical
he de ising o a ield- easible and ime-e ec i e decon-
amina ion p o ocol (See de ails in Sec ion 4).
2. Con amina ion Con ol P ac ices o Spacec a
Ha dwa e
Du ing he ARADS campaigns, we ollowed s a e-o - he-
a con amina ion con ol p ocedu es ypical o hose o
p epa e o Ma s landed missions. The modi ica ions i he
equi emen s posed by he e es ial ield ial. Ca osso
(2005) o e ed comp ehensi e backg ound in o ma ion
on spacec a decon amina ion and e i ica ion echniques
sha ed wi h ecen missions a ge ing plane a y samples.
Examples include he 2007 Phoenix Mission and o he
missions o Ma s such as he 2011 Ma s Science Labo a o y
(MSL) (Maha y e al., 2004) and he 2020 missions cen-
e ed on he analysis o molecula o ganic compounds, and
he 2016 OSIRIS Rex o he ca bonaceous nea -Ea h
as e oid Bennu (Lau e a e al., 2017; Dwo kin e al., 2018).
Al hough hese missions we e expec ed o ad ance
decon amina ion p ocedu es o sampling and sample han-
dling mechanisms, hey ha e ye o p og ess beyond he
2007 Phoenix Lande expe ience due o cos caps and slow
ad ances in ma e ial echnology (Mille e al., 2008). The
Phoenix Mission, ope a ed on Ma s om 2008 o 2009,
o e s s a e-o - he-a con amina ion con ol o spacec a
ha dwa e analog o ARADS.
The Lande ’s obo ic a m-moun ed digging scoop o
ISAD collec ed and deli e ed ice-bea ing egoli h samples
o he The mal E ol ed Gas Analyze (TEGA) o de e mine
indica o s o habi able condi ions in Ma s ice-cemen ed
g ound, equi ing s ic con amina ion con ol (10 ppb) on
he digging scoop o p o ec he scien i ic esul s om Ma s.
Once assembled, he compa men alized scoop su -
aces we e inaccessible o cleaning, challenging c oss-
con amina ion con ol be ween samples scooped om he
ma ian soil. Unlike he 1975–1976 NASA’s Viking lande s,
which we e hea ea ed in hei en i e y (e.g., So en and
Snyde , 1976), he Phoenix Mission sampling chain (blades
and scoop) main ained only low o ganic con amina ion Le el
75 A/10 equi emen , ha is, 100 ng/cm
2
(Bo son, 2005).
Con ol and assessmen o con amina ion on he Phoenix
TEGA included acuum bake-ou /d y-hea s e iliza ion,
con ainmen o anspo and es ing, and con amina ion
cha ac e iza ion on an enginee ing mock-up ligh model.
Du ing Assembly, Tes , and Launch Ope a ions (ATLO)
ligh ha dwa e in an ISO 14644-1 Class 8 con olled en-
i onmen , biobu den equi emen is £10 CFU/m
3
(ai
samples), 2000 CFU/m
2
(su aces), and £1 CFU/
5- inge glo e p in (ECSS, 2008a, b). Du ing asep ic op-
e a ions (Class <7), ha dwa e assembly occu s on lamina
low unde a s ic e NASA S anda d assay (NSA)-based
biobu den le el o <1 CFU/m
3
(ai ), £300–400 CFU/m
2
(su aces), and <1 CFU (pe glo ep in ).
Assuming ha he NSA can de ec only 1% o he o al
iable and non- iable bu den, 400 CFU/m
2
co esponds o
he h eshold o asep ic ope a ions o £40,000 mic obial
cells/m
2
(o £3–4 cells/cm
2
) achie able by ARADS.
3. Sou ces o Con amina ion
Whe he o no a o eign ma e ial is a ‘‘con aminan ’’
depends on he objec i e o he planned analysis and only i
he ma e ial in e e es wi h he ins umen s’ measu emen .
Fo eign cellula ma e ials and o ganic bioma ke s de ec ed
in ARADS co es and ha dwa e a e con aminan s. ARADS
con amina ion con ol (Sec ion 4) a ge ed an h opogenic,
en i onmen al, and c oss-con amina ions while add essing
hei isk le el be o e and a e mi iga ion.
(1) An h opogenic con amina ion by human- ended
au oma ed d illing es s is una oidable and a signi i-
can conce n o biologically deple ed e es ial and
ma ian en i onmen s. In A acama’s biologically lean
su ace en i onmen , ai bo ne mic oo ganisms a e
lesse conce ning due o ai low dispe sion. Con-
e sely, an h opogenic mic obiome con amina ion o
soil and ai ia handling, clo hing, and skin pa icles
du ing daily ope a ions can comp omise esul s om
he li e-de ec ion sui e.
(2) En i onmen al con amina ion is a unique p oblem in
e es ial ials, wi h h ee po en ial sou ce en ies
in o he sample pa h: (i) non-pu a i e biological
( iable spo es, mic obes, cellula emains); (ii)
ossil molecula o ganics de ec able by LITMS and
SOLID; and (iii) ai bo ne mine al dus and sal s
de ec able by he we chemis y ins umen . The e is
a simila oppo uni y o ai bo ne ino ganic con-
amina ion on he ma ian su ace, which is an un-
likely sou ce o biological con aminan s. Ai bo ne
dus om he as oligo ophic dese pa emen o
d ill si es was ini ially no o g ea conce n wi h
bio ba ie s o p o ec sample-handling elemen s
when no in use. S ill, ai bo ne biology esul ed in
an unexpec ed po en ial con aminan o ARADS
(Sec ion 6.2).
(3) C oss-con amina ion could occu be ween di e en
sedimen a y s a a d illed as powde ed samples and
mixed om he a ge dep h; once in he scoop, a new
sample could mix wi h sedimen esidue adhe ing o
he inside om p e iously d illed in e als.
4. Me hods o Con amina ion Con ol
We desc ibe by-design, ope a ional, and con amina ion
con ol echniques we es ed in ARADS.
4.1. Con amina ion emedia ion by design and
ope a ional me hods
To emedia e c oss-con amina ion, we used s ainless-s eel
o aluminum sampling ha dwa e, a oided so ma e ials
and polyme s poo ly conduci e o ield decon amina-
ion, and chose single-use p e-cleaned s ainless s eel un-
nels and clean-ce i ied bo osilica e glass ja s. The sample
acquisi ion and handling sys em design also enabled con-
aminan dilu ion by accep ing a ela i ely la ge sample size,
up o 60 g, compa ed wi h he expec ed esidual c oss-
con aminan s (Fig. 12).
The gene al con amina ion emedia ion ope a ions inclu-
ded p o ec i e shields usage and d ill ope a o s aining
guidelines ha in ol ed: (1) mind ul walking in
ARADS CONTAMINATION CONTROL STRATEGY IMPLEMENTATION 1307

app oaching he o e downwind o a oid s i ing dus
om he playa pa emen (Fig. 3J); (2) only wo ope a o s
ending he d ill in-sim (Fig. 3H); (3) use o acemasks
and sani ized glo es (e hanol [E -OH]-wiped) when ap-
p oaching ha dwa e a close con ac wi h samples o
wo king on he ins umen s ou le s; (4) hand-sani iza ion
be o e wea ing glo es; (5) ouch no c i ical equipmen
su aces and ools o con ac wi h sampling su aces wi h
un-sani ized o con amina ed glo es; (6) a oid con ac
be ween he clean glo es and body’s su ace, o any
comp omised su ace; (7) use o clean/s e ilized ools
whene e possible; and (8) wea a p o ec ion sui when
app op ia e (Fig. 3F).
We used p o ec i e media o p oac i e con amina ion
con ol be o e, du ing, and be ween ope a ions. Fo ins-
ance, be o e assembly on he o e deck, we anspo ed
double-w apped cleaned unnels in sani ized plas ic con-
aine s (S e ili e TM) wiped down wi h high-pu i y iso-
p opyl alcohol (IPA). While he o e mo ed owa d
he d illing si e, we p o ec ed he cleaned d ill, scoop,
and unnels wi h hea y-du y Ul a High Vacuum (UHV)
Aluminum Foil (AllFoil, Inc.), which was emo ed only
be o e use.
Be ween each d illing cycle, we kep he bio-ba ie in
place un il he d ill was eady o ope a e (Fig. 4C), he scoop
was abou o accep d illed samples, and he unnels could
ecei e and ans e samples o analysis. We main ained
a con amina ion- ee sample pa h o each li e de ec ion
ins umen . Fo ins ance, he SOLID’s inle al e opened
only be o e he sample’s elease in o he analy ical ool
(Pa o e al., 2008).
A he end o each Sol, he o e -moun ed d ill s ing
and scoop we e insed o wi h dis illed wa e (DW),
w apped in Al oil, and le onsi e o e nigh . The unnels
we e unclamped om he o e deck, w apped in Al oil,
and anspo ed o he Yungay S a ion (La . 244¢53.05’’S,
Long. 6955¢9.51’’W) o a new cleaning cycle be o e he
successi e Sol’s ope a ions. Each day, he o e -moun ed
d ill s ing was ‘‘ ouched up’’ and cleaned (done on p e i-
ously deep-cleaned su aces) be o e use.
4.2. Chemical cleaning and disin ec ion
We es ed he deep-cleaning e ec i eness on wo Ice-
b eake d ill segmen s: A (d ill ip, 0–25 cm) and C (40–
60 cm) in Tes 2 (Ma ch) and segmen B (20–40 cm) in
Tes 3 (Sep embe ) oge he wi h he sample handling
ha dwa e elemen s (Tes 4). We used chemical p ecision
cleaning, a co e componen o bo h Quali y Assu ance o
obo ic landed missions o Ma s and PP (e.g., F ick e al.,
2014).
In 2019, we did no use hea s e iliza ion because su -
aces could be easily e-con amina ed in he ield, and
decon amina ion cycles would equi e addi ional ime.
Fu he , hea s e iliza ion would no emo e o ganic con-
aminan s. Tables 1 and 2 summa ize he chemical cleaning
FIG. 3. (A) Disassembled d ill s ing, scoop,
and unnel p epa ed o ield deep cleaning
and es ing; (B) p elimina y s ep (dis illed
wa e inse) o emo ing pa icula e ma e ial
om he d ill s ing; (C) ATP swab assay o
one ins umen ’s unnel ha dwa e; (D) Su ace
Swab Assay samples o SOLID co-analysis;
(E) ATP molecule (le ) and hand-held lu-
minome e wi h liquid (le ) and su ace
( igh ) sampling de ices. ATP, adenosine i-
phospha e.
1308 BONACCORSI ET AL.
and mic obial educ ion s eps we applied; hei speci ic
e ec i eness o ARADS is add essed in Sec ion 5 and
Figs. 6A–H and 7.
4.2.1. Cleaning and disin ec ing agen s. The p ecision
cleaning in ol ed (1) adenosine iphospha e (ATP)- ee
dis illed wa e (DW) o liquid ch oma og aphy/mass spec-
ome y (LC/MS) g ade wa e o emo al o isible di
(adhe ing clays and dus pa icles); (2) sol en s wi h de-
c easing pola i y (pu e g ade ace one, me hanol, and E -OH)
o emo e he a oma ic and alipha ic molecula o ganics;
and (3) mic obial educing agen s, o example, 60–80% E -
OH, high-pu i y IPA, 0.5–5% household bleach (NaClO;
esh ba ch, unopened bo le as di ec sunligh and o ganic
ma e inac i a e NaClO), and 3% hyd ogen pe oxide
(H
2
O
2
) o dis up ing bac e ial bio ilms and o inac i a e/kill
mic oo ganisms.
NaClO and H
2
O
2
we e applied indi idually o simul a-
neously. We used a ious dispensing me hods and mul iple
applica ions o maximize he chemical exposu e o he a ge
ha dwa e, o example, bleach-soaked (imp egna ed) poly-
p opylene wipes w apped a ound he s ing’s su ace, wi h
w aps molded a ound he d ill’s spi als o 1 h. A e wa d,
he d ill was wiped o , ai -d ied, and ho oughly insed wi h
ca. 0.5–1 L o ATP- ee DW.
4.2.2. Dispensing and wiping. We adap ed echniques
based on ha dwa e opog aphy; in some cases, we used p o-
essional sp ay bo les o mis , sp ay, and s eam each che-
mical. S ill, highly ola ile sol en s e apo a e quickly om
he a ge ha dwa e, dec easing hei cleaning e ec i e-
ness. In some cases (d ill deep cleaning), we p e e ed
s oke-wiping ha dwa e wi h lin - ee anhyd ous sol en -
imp egna ed wipes o emo e con aminan s ins ead o di ec
sp aying on o he ha dwa e.
The s anda d Quali y Assu ance o obo ic landed mis-
sions o Ma s equi es pe iodic p ecision cleaning o expo-
sed ligh ha dwa e su aces wi h high-pu i y IPA-soaked
lin - ee swabs and wipes (e.g., Ca osso, 2005). We im-
p egna ed wipes (by sp aying su icien eagen s on o hem)
o he mos e icien disin ec ion (Panousi e al., 2009).
In o he cases (e.g., unnel’s na ow ou le s), we nebu-
lized aqueous IPA se e al imes o maximize exposu e o
he disin ec ing agen . Finally, a las i e a ion wi h (IPA)-
soaked low-lin Kimwipes (KIMTECH KimWipes Task
Wipe s) emo ed cellula emains om he su ace o max-
imum disin ec ion. The wiping bes p ac ice we applied
in ol ed consis en p essu e on e e y s oke, wo king om
op o bo om ( o he d ill s ing) and ‘‘pull and li ’’ om
he in e io (cleanes ) o he ex e io (di ies ) a eas ( o
scoop and unnels).
FIG. 3. (Con inued) (F) Mi iga-
ion con amina ion du ing ARADS
2016 g ound u h sampling o soil
pi s (Pho o c edi : NASA Ames); (G)
con ainmen se up a Yungay Base-
camp be o e ield- es ing o he
NASA Godda d’s Ion T ap Mass
Spec ome e . (H–K) ARADS p ac-
ices o con amina ion mi iga ion
examples. See desc ip ion in Sec ion
4.1. (H) Reduced ield pe sonnel
du ing o e ops; (J) minimized un-
necessa y walking on playa owa d
o e s; and moni o ed oo s eps on
g ound si es o scien i ic in e es , ha
is, pos - lood desicca ed bio ilms (K).
ARADS CONTAMINATION CONTROL STRATEGY IMPLEMENTATION 1309
When olded, he wipes p o ide up o eigh clean, us-
able sides o p e en e-con amina ion ac oss he cleaned
su ace.
4.2.3. Wa e inse. A hi d s ep included ho oughly
insing wi h a s eam o ATP- ee gas ch oma og aph–mass
spec ome e (GC-MS) g ade wa e o he comple e emo-
al o cleaning agen s, any Kimwipes’ ibe esidue, dead/
inac i a ed mic obes, and cellula cons i uen s.
4.3. Ha dwa e con amina ion e i ica ion assays
We used wo independen con amina ion e i ica ion
me hods: he ATP luminome y assay o iable o al bio-
bu den and he Fluo escence Sandwich Mic oa ay
FIG. 4. (A) Ou -o -sim biobu den con amina ion es s wi h he TRIDENT d ill in wo king posi ion; (B) Top: Iceb eake
d ill segmen including he D ill bi (0–25 cm). Bo om: d ill Segmen 20–40 cm wi h a smoo he su ace inish han he
D ill bi ; (C) moun ed d ill a e a 24-h exposu e o he GP en i onmen and Al oil bio-ba ie p o ec ing d ill be o e ield
es ing; (D) po en ial sou ces o en i onmen al con amina ion om su ace biology a he GP Si e: Vascula plan biomass;
and (E) ield o cyanobac e ia-colonized hali e. TRIDENT, The Regoli h and Ice D ill o Explo a ion o New Te ains.
Table 1. Mul i-S ep P o ocol, as Desc ibed in Table 2
These s eps in ol e mechanical cleaning (1–5), sol en -based molecula o ganics emo al (6–8), physical and chemical mic obial
educ ion/s e iliza ion (9–15), and p e en i e ac ions (16–17). Household bleach is 5.25% sodium hypochlo i e (NaClO).
DW =dis illed/deionized wa e ; E -OH =e hanol; IPA =isop opyl alcohol; LC/MS =liquid ch oma og aphy/mass spec ome y; Me -
OH =me hanol.
1310 BONACCORSI ET AL.
Immunoassay (FSMI) o b oade cha ac e iza ion o mi-
c obial axa and hei molecula and cellula con aminan s.
Bo h echniques, desc ibed nex , in ol e a sol en swab
assay o he pos -cleaning ex e io d ill s ing segmen s and
he in e io su ace o he scoop and unnel.
Bes p ac ices include a consis en swabbing echnique
(ho izon al, e ical, and diagonal), di ec ional p essu e, and
ime ca. 5 min (see e.g., Ba engol z, 2005). Swab samples
equi e subsequen we ex ac ion o con aminan s analysis.
Depending on he su ace oughness, we used wo swabs o
e ie e he ATP om he ha dwa e su ace and combined
he esul ing alues o assayed smalle su aces i co e ed
by mine al dus .
4.3.1. ATP luminome y assay. We elied on he
ATP Swab assay o he in si u eal- ime, p e-sc eening
con amina ion isk- educ ion and disin ec ion ce i ica ion
because all li ing o ganisms use ATP as he uni e sal en-
e gy ca ie (Lundin and Tho e, 1975a, 1975b), and his
molecule quickly deg ades a e cellula b eakdown (Holm-
Hansen and Boo h, 1966).
The assay is used o he biobu den moni o ing o
spacec a ha dwa e in clean ooms (NASA P ocedu al
Requi emen s 8020.12D; 2011, Venka eswa an e al.,
2003). The Hygiena ATP assay sys em (Fig. 3E) includes
a sel -calib a ing (25–28C) handheld Luminome e (En-
SURE) wi h a dynamic ange o 0.1–2000 em omoles,
ope a ing wi h an ul asensi i e ATP su ace swab (Supe -
Snap), whose limi o de ec ion (LOD) is 0.1 ·10
-15
moles,
o 0.1 moles ATP.
The luminome e measu es he ligh emi ed by he i e-
ly’s (Pho inus py alis) luci e in-luci e ase (L-L) enzyma ic
eac ions (pH 7.6-bu e ed) binding wi h he ATP eleased
by li ing cells in he p esence o oxygen and magnesium
(Balkwill e al., 1988), p opo ionally o he ATP in he
sample (Lundin and Tho e, 1975a; McEl oy and DeLuca,
1983). The pho ode ec o con e s he gene a ed pho-
ons in o Rela i e Luminosi y Uni s, o RLUs (15-s ead-
ings), ha can ansla e in o ATP concen a ions (1 RLU
equi alen o 0.1 moles ATP) wi h dilu ions o ATP
s anda d (Na_ATP sal ).
ATP þDLuci e in þO2þLuci e ase Mg2þ

>Oxyluci e in þAMP þPy ophospha e þCO2
þh (560 nm)
4.3.2. Fluo escence sandwich mic oa ay immunoassay.
The Quali y Assu ance o obo ic landed missions o Ma s
(Phoenix, MSL, and 2020) equi es ollow-up s anda d
molecula cleaning e i ica ion by Sol en Swab Analysis
(SWA) o ligh ha dwa e’s swabbed su aces wi h a F eon-
based sol en ia Fou ie ans o m in a ed (FTIR) spec-
oscopy o GC-MS (Ande son e al., 2002; Maha y e al.,
2004; Blakkolb e al., 2014).
A subse o pos -cleaned ha dwa e (Sol 5 and Sol 6) ana-
lyzed o ATP (Table 6) was also co-analyzed epu posing
he SWA o use by he SOLID’s FSMI a ge ing SOLID-
ele an biological con amina ion in place o con amina ing
compounds de ec able by FTIR o GC-MS. We used Te lon
Table 2. Con amina ion Con ol P ac ices Used o he Ro e As obiology D illing S udies Subsys ems
See Table 1 o an explana ion o each p o ocol’s coded i ems.
SHTS =Sample Handling and T ans e Sys em.
ARADS CONTAMINATION CONTROL STRATEGY IMPLEMENTATION 1311
analyzed wi h he LDChip200 immunosenso o b oade
cha ac e iza ion o he Sol 5 and Sol 6 ha dwa e backg ound
esidue (ATP syn hase and cellula p o ein bioma ke s). The
ull da ase and immunogens’ in ensi y peaks a e p o ided
in Supplemen a y In o ma ion Fig. S1, Table S2a and S2b.
We could pe o m only one sample ex ac ion and con-
cen a e he olume o imp o e de ec ion. In ou expe ience,
no di e ences in luo escence alues esul ed when using
he same ex ac wice. E o ba s co espond o he s anda d
de ia ions o h ee in e nal eplica es o each an ibody a ay
we es ed.
5.2.2.1. ATP syn hase an ibodies. ATP syn hase subuni 1
(ASB and 20_p o ASF1 [ATP syn hase subuni 2]) is a
p o ein ha o ms he in acellula ene gy s o age molecule
ATP in he mi ochond ia. We de ec ed ATP syn hase in all
ha dwa e su aces co-analyzed wi h he ATP Luminome y
Assay (Table 6). Al hough he signal in he blank swab
(nega i e con ol) was high, a e sub ac ing he nega i e
con ol, he immunoassay yielded nea -ze o FI signals
o ATP syn hase (20_p o ASF1) in he scoop and d ill
(Sep embe 20 assay).
The only FI signal o he p o ein’s o he subuni ASB
(FI: 2347 –850) ela es o Sample 1 (scoop, Sep embe 19)
and subuni ASF1 in ai bo ne samples 9 and 10, in ag ee-
men wi h alues o ATP abo e 6.0 RLUs (Table 6). High
RLU luminescence ma ches he inc eased luo escence
signal o ASB 2, whe eas low RLU co esponds o nega i e
de ec ion (FI: 0) o ace signals (FI: 55–235). The wo
signals appea posi i ely co ela ed (R=0.836) when inc-
luding he wo ai samples, bu he co ela ion d ops when
conside ing only ha dwa e samples (N=5), and mo e anal-
ysis is necessa y o a obus co ela ion.
5.2.2.2. Cellula and p o ein bioma ke an ibodies. The
FI peaks (max alues) agains he mic obial axa and
molecula bioma ke s de ec ed by he LDChip200 we e
highe han hose agains ATP syn hase in he pos -cleaned
ha dwa e (Table 7 and Fig. 8). Fu he , we compa ed each
bioma ke de ec ed in he ha dwa e wi h hose iden i ied in
he ai dus samples (Supplemen a y Table S1a, b).
The compa ison in o ms whe he he backg ound con am-
inan was om an ai bo ne sou ce o po en ial c oss-
con amina ion om d illed sedimen s esidues ans e ed o
he sample handling ha dwa e and analysis. Fi y pe cen o
less o axa ( ace esidues) de ec ed in he d ill, scoop, and
unnels we e also in he dus . In compa ison, mo e han 50%
we e de ec ed only in he backg ound esidue ans e ed om
d ill o unnel o scoop (pu a i e subsu ace biome).
Ve y low signals (FI <400–600) agains Anabaena sp. a e
ubiqui ous in pos -cleaned ha dwa e and dus samples. The
highes FI-posi i e alues in he swabbed ha dwa e sam-
ples a e agains p o eins G am-nega i e bac e ia ances al
be a-lac amase (GNCA), Gammap o eobac e ial ances al
be a-lac amase (GPBCA), and Ni S2 (Lep ospi illum e -
ooxidans). The pep idoglycan (polyme suga s and amino
acids o ming bac e ia cell wall) is ound only in he 19-Sep
scoop (FI <800 and 8.4 –1.6 amoles ATP/cm
2
) bu no in
he dus . Ve y weak signals agains Bacillus sub ilis and
S ep omyces sp. a e de ec ed only in pos -cleaned ha dwa e
om Sol 6 (Sep embe 20), whe eas he peak in ensi y
(<700) is agains Planococcus sp., a G am-posi i e psy-
ch o ole an , halophilic bac e ium.
FIG. 8. Linea plo wi h nega i e co ela ion (R=-0.673)
be ween pos -cleaning su ace ATP backg ound (ATP
luminescence) and Immunoassay-based mic obial axa
backg ound (Fluo escence max in ensi y) o Sol 5 and Sol 6
ha dwa e. No e he weak signals (FI: <700), excep o he
d ill bi ’s posi i e signal (FI >1000), as de ailed in Table 7.
FI, luo escence in ensi y.
Table 7. Pos -Cleaning Mic obial Cells and P o ein Bioma ke s De ec ed by he LDChip_200
in he Ha dwa e Backg ound and Co-Analyzed o Adenosine T iphospha e Bioma ke
by Adenosine T iphospha e Luminome y (Thick-Bo de ed Columns)
F om le o igh : o al ATP swab Assay o in-sim cleaned HD; ATP pe su ace uni ; maximum alue o FI (peak) agains mic obial axa
(whole cell) and molecula bioma ke s (p o eins). ‘‘P esence in ai sample s’’ indica es he ac ion (%) o mic obial axa and p o ein
bioma ke s de ec ed in one o bo h ai bo ne sample s and he HD backg ound. We calcula ed he pe cen o molecula bioma ke s in ai
sample s om he an ibodies aised agains whole cells, p o eins, bio ilms, and sedimen mix u e cul u es in he HD esidue (Supplemen a y
Table S2a and S2b) and hose de ec ed in he wo ai sample s (Table 9).
GNCA =g am-nega i e bac e ia ances al be a lac amase.
1318 BONACCORSI ET AL.

The su ace swab o he pos -cleaned ha dwa e yielded
ela i ely low backg ound con aminan s (Table 7), indica ed
by aces (<300) o weak (<1000) FI signals, excep o a ew
mode a e signals (1000–3000). The in ensi y signals om
p o ein bioma ke s con aminan s (FI peak *1000–1600) a e
also highe han hose agains cellula ma e ials (FI peak:
*300–1000). A ew high signals (>3000) pe ained only o
ma e ials om he ai sample s (Fig. 10 and Table 9).
The esul s sugges ha sol en -cleaning and mic obial-
educ ion s eps e ec i ely mi iga ed esidual and ac i e
biobu den con amina ion in he ARADS ha dwa e, wi h a
ew excep ions. The chip de ec ed a mode a e FI signal
agains Geobac e sp. (FI: 1008) and he Gammap o eo-
bac e ial lac amase p o ein (FI: 1472) in he pos -cleaned
d ill bi (Sol 6, Sep embe 20).
Howe e , his highes backg ound in ensi y signal co e-
sponds o one o he lowes signals de ec ed agains ATP
syn hase (FI: <300), which co ela es wi h he ATP bio-
ma ke (1.2 –0.6 amoles/cm
2
). The ATP backg ound con-
amina ion was equi alen o aces o iable biobu den o
ca. £1 cell/cm
2
, o a o al o *100–200 cells in he d ill bi
(0.6 –0.0 amoles/cm
2
ATP) based on *1–2 amoles ATP
pe bac e ial cell (Okanojo e al., 2017).
Fu he , a nega i e co ela ion exis s be ween he FI peak
agains mic obial cells (g ay columns) and he ATP back-
g ound signal ( =-0.673) as shown in Fig. 8. The ela-
ionship sugges s ha only a negligible ac ion (i any) o
he esidual mic obes de ec ed by he LDChip200 may be
iable (ATP ac i i y) and ha he FI ela es o esidual
aces o dead o pa ially agmen ed cells. The e is no
signi ican co ela ion be ween ATP backg ound and FI
peak agains p o eins ( =-0.196), whe eas no co ela ion
exis s be ween he cells’ and he p o eins’ FI peak
( =-0.017). Co ela ion da a a e no p esen ed.
5.3. Ai bo ne con amina ion
Figu e 9A–D and Table 8 illus a e moni o ing se up o
ai bo ne biomass lux o collec o s 1 and 2.
5.3.1. An h opogenic and en i onmen al ai bo ne bio-
mass. The ATP-based ai bo ne biomass lux a Collec o 1
anged om 23.7 amoles o 47.4 amoles/cm
2
ac oss he
6-Sol simula ion. The deposi ion a es o an h opogenic
ATP a basecamp, a e age o 217.8 moles/(m
2
$day), we e
4.2 imes highe han he a es o en i onmen al ATP, which
a e aged 51.6 moles/(m
2
$day).
The lux o ai bo ne an h opogenic deli e ed o e 5 days a
basecamp (Coll. 1) was 110.7 amoles, a e aging 22.1 amoles/
(cm
2
$day), o 221 moles/(m
2
$day), whe eas Playa Collec o
2 ecei ed a o al amoun o 46.4 amoles/cm
2
ATP o e he
same pe iod exposu e. A e backg ound sub ac ion (Bkg :
20.6 amoles/cm
2
), he o al lux o ai bo ne en i onmen al
biomass o he playa (Coll. 2) was 25.8 amoles/cm
2
, equi a-
len o 5.2 amoles/(cm
2
$day) o 51.6 moles ATP/(m
2
$day).
Fu he , by assuming ha basecamp Collec o 1 would
ecei e ai bo ne dus in simila amoun s o he Playa Col-
lec o 2, he heo e ical ne an h opogenic componen could
be calcula ed as he di e ence be ween he wo ai bo ne
luxes a he wo collec o s (Coll 1 Flux -Coll 2 lux),
which is 166.2 moles ATP/(m
2
$day).
Table 8 epo s he calcula ed ai bo ne lux (en i on-
men al biomass) based on ATP swab da a and no malized
FIG. 9. (A, C) Ai bo ne collec o s’ loca ion and deploymen . (A) Basecamp Collec o 1 ou side he Science aile (da k
ed); (B) Collec o 2, wi h basecamp in he backg ound; (C) Top iew o he 23 ·23 cm collec ing su ace and sub-sampling
a eas (A–H). Supplemen a y Tables S1a and b in he Supplemen a y In o ma ion epo he su ace a ea and ATP swab da a
used o lux calcula ions. (D) Moni o ed 6-Sol ai bo ne con amina ion in collec o s 1 and 2 as ATP a omoles/cm
2
.
Es ima ed daily lux da a a e om Table 8. Squa e symbols and e o ba s indica e he a e age and STD (Column F,
Table 8) o indi idual measu emen s (no shown he e; see he g ay-shaded Column J in he Supplemen a y Table S2
ins ead). The solid ba s indica e he o al amoun o su ace ATP measu ed o e 5 days (Column D). STD, s anda d
de ia ion.
ARADS CONTAMINATION CONTROL STRATEGY IMPLEMENTATION 1319
o he en i e collec o ’s su ace (23 ·23 cm). Fo collec-
o s 1 and 2 ai bo ne lux calcula ions, see Supplemen a y
In o ma ion o p e-deploymen se ups, pos -sani a ion ATP
backg ound de e mina ion (Supplemen a y Table S0a and
b), and daily ai bo ne biomass aw da a (Supplemen a y
Table S1a, b and Fig. S3).
5.3.2. Mic obial axa in ai bo ne samples. The wo col-
lec o s we e loca ed downwind om he Playa and ecei ed
a highe lux o con aminan s om he wind-scou ed playa
su ace sedimen s (Fig. 13 [3, 4]). LDChip200 de ec ed in
si u bac e ia, a chaea, and molecula bioma ke s a he wo
si es, wi h mos ly e y weak signals o he ela i e FI (FI
<600–1000) ac oss he wo si es, bu a ew excep ions
(Table 9 and Fig. 10). The e a e a ew in ensi y signals
(weak signals) in bo h Basecamp Collec o 1 (sample #9)
and he dis al allu ial an/Playa Collec o 2 (Sample #8),
al hough #9 has mo e posi i e ones.
The 10 m ack sample is a li le mo e con amina ed gi en he
mode a e signals (FI >1000) o an ibodies agains De-
chlo omonas,Desul o ib io (Coll 1); he highes luo escence
peak (FI >6000) ela ed o he PhaC1 (a p o ein om aqua ic
cyanobac e ia, Anabaena sp., Nos oc sp.) and he Fe idoxin
enzyme. The aqua ic mic o lo a was p esen in bo h collec o s,
bu he cyanobac e ium Xenococcus, cha ac e is ic o semi-a id
hypoli hic communi ies ( he A acama B sequence, Wa en-
Rhodes e al., 2006), occu ed only in he dis al Collec o 2.
O e all, he posi i e immuno- eac ions occu ed wi h an-
ibodies aised agains : (1) G am-nega i e acidophilic i on- and
sul u -oxidizing bac e ia (Acidi hiobacillus,Desul o ib io); (2)
pe chlo a e- educing Be ap o eobac e ia (Dechlo omonas,
Ideonella), Gammap o eobac e ia (Shewanella,Pseudomonas,
Gammap o eobac e ia ances al be a-lac amase); (3) Cyano-
bac e ia: aqua ic (Anabaena,Nos oc) and hypoli hic (Xeno-
coccus); (4) G am-posi i e Ac inobac e ia (S ep omyces)and
Fi micu es (Bacillus,Desul ospo osinus); (5) s ic ly anae obic
sul a e- and me al- educe s bac e ia (Desul ospo osinus,She-
wanella); and (6) halophilic he e o ophic a chaea and me-
hanobac e ia (Me hanobac e ium).
The immunog am da a in Table 9 show ha he mic obes
exclusi ely p esen a Collec o 2 a e G am-nega i e bac e ia,
wi h G am-posi i e (Fi micu es, ac inobac e ia) and A chaea
in he basecamp’s ai sample . The immunoassay da a a e no
s a is ically ele an (only a collec o o each a ea), whe eas
he immunog ams ma ch well he ATP da a.
5.4. Mic obial biomass o d illed and su ace sedimen
Figu e 11 shows he downco e dis ibu ion o ATP bio-
ma ke om h ee Playa d illing si es. The a e age ATP
bioma ke is 100–250 moles/g and appea s o dec ease wi h
dep h in Hole 5 (60 o <0.2 moles/g, below de ec ion limi
[DL]) and Hole 2 ( om 350 o 5 moles/g).
The dis ibu ion o ATP bioma ke s in hese co es app-
ea s o be om complex pu a i e li e (e.g., Wa en-Rhodes
e al., 2019) a he han om con aminan s ans e ed
du ing d illing ia ha dwa e, pos -d illing sample manipu-
la ion (e.g., hand skin eading 200 moles), o ai bo ne
biomass- ich sedimen nea by (Fig. 13), wi h hese con-
aminan s being p e en ed by ou s ic con amina ion
con ol p ac ices du ing ope a ions.
Table 8. Flux o Ai bo ne En i onmen al (Collec o 2) and An h opogenic
(Collec o 1) Biomass
F om le o igh : Column B: Swabbed su ace a ea ou o he o al collec o su ace (529 cm
2
); Column
C: ATP swab alues no malized o 529 cm
2
; Column D: ai bo ne biomass in a omoles ATP pe su ace uni
(amol cm
2
). Columns E and F: Summa y s a is ics (min, max, A g., and STD) o da a ela i e o indi idual
su aces sub-a eas A, B, C, D, e c. (no included, see Table S1 in he Supplemen a y In o ma ion). The las wo
ows epo a e aged ATP lux as em omoles pe squa e me e pe day [ moles/(m
2
$day)] and a omole/
(cm
2
$day).
1320 BONACCORSI ET AL.
FIG. 10. Immunog am wi h he ela i e FI o he LDChip200 posi i e de ec ion o mic oo ganisms and polyme ic biological ma ke s a he basecamp and dis al
collec o s. No e ha some immunogens a e p esen only in ei he one o he wo collec o s, whe eas o he s occu in bo h ones. See Sec ion 5.3.2. o a desc ip ion and
Table 9 o he lis o an ibody names.
1321
The unusually highe alues o ATP in he uppe -le el
samples (0–10, 40–50, and 10–20 cm) and he dec eases o ATP
wi h he bo ehole dep h (50–60, 60–70, and 30–40 cm) migh be
due o na u ally occu ing me abolically ac i e biomass p esen
in hese le els as he esul o he la es ponding e en s.
Fo compa ison, 15.1–32.4 ng/g, o ppb, o o al amino
acids (Leu, Val, Se , Ala, and Gly) we e de ec ed by he
MILA ins umen in h ee subsu ace samples acqui ed in
2019 ( alues calcula ed om Mo a e al., 2020; Supple-
men a y Table S2).
Finally, ‘‘G een Pa o ’’ d illed da k b own clays (Ma ch
2019) yielded highe alues (*2·10
3
o *7·10
4
cells/g)
agains he coa se-g ained su ace pa emen (*10
2
–10
3
cells/g), wi h no eco e y o mic obial ATP in i e ou o
FIG. 11. Dep h dis ibu ion o mic o-
bial ATP in h ee holes exp essed as
moles o ATP bioma ke pe g am o
sedimen . Ho izon al e o is STD
(Table S3 in he Supplemen a y In-
o ma ion).
Table 9. Peak Numbe s, An ibody Names, and Immunogens o In ensi y Peaks
o he Immunog am o Ai Samples om Fig. 10
1322 BONACCORSI ET AL.
se en ials (Supplemen a y Table S3). Supplemen a y
Figu e S2A–E epo s p o ocol es esul s, and Supple-
men a y Table S3 p o ides Summa y s a is ics o he
ARADS su ace and d ill samples da ase .
5.4.1. Su ace dis ibu ion o biomass con aminan a
Playa d ill si e. We measu ed he su ace dis ibu ion o
o al, ee, and mic obial ATP in smec i e clay- ich playa
sedimen s as po en ial con aminan indica o s o he ARADS
si es (Fig. 12 and Supplemen a y Table S3 in he Supple-
men a y In o ma ion). Samples included (1) a 3 mm laye o
he desicca ed mic obial ma as posi i e con ol, ha is,
4.7 –0.58 ·10
4
moles o al ATP/g (N=9); (2) exca a ed clay
om a nea by pi (3–5 cm-dep h), ha is, 7.2 –0.7 ·10
3
moles/g (N=10); and (3) ne e -ponded su ace clay (0–
3 cm-dep h), ha is, 212 –268 moles/g (N=6).
These alues a e equi alen o an es ima ed iable
mic obial biomass o 10
3
cells/g (0–3 cm-dep h) and 10
5
cells/g (3–5 cm-dep h) o e y high le els (10
6
cells/g) in he
3 mm laye o he d ied mic obial ma (posi i e con ol
samples) a 4-log o a 5-log highe mic obial biomass han in
d illed sedimen s o he majo i y o he Playa su ace.
This unexpec edly highly ac i e su ace biomass ela ed
o a hin c us o desicca ed mic obial ma (Fig. 3K) we
sampled om small a eas o he pos - looded Playa and was
s ill p ese ed a e a sho -li ed pond o med a i s ma -
gin du ing an ex eme lood e en in 2017 (Pe sonal com-
munica ion; Kim Wa en-Rhodes, 2019), 2 yea s be o e ou
campaign. In epheme al ponds, spo e- o ming cyanobac-
e ia can p oli e a e and lea e pos - looded desicca ed ye
iable biomass on he su ace.
The desicca ed, e y b i le bio ilm could be e oded,
ca ied, and deli e ed by wind, inc easing he con amina ion
isk when en aining exposed co ed ma e ial o he scoop
and unnels du ing he sample ans e (Fig. 13). Howe e ,
mos su ace biomass was 3- o a 4-log lowe han bo eholes
sedimen s o he high biomass pa ches.
5.5. Con amina ion ans e o d illed samples
An in eg al componen o he con amina ion plan included
a sou ce- o-sink assessmen o po en ial con amina ion in he
d illed samples (ha dwa e- o-sample ans e , ai bo ne bio-
mass con amina ion on sampling ha dwa e, e c.). Resul s
om hese asks a e in used in o a concep ual model (Fig. 13)
and in eg a ed in o he Discussion (Sec ion 6).
Figu e 12 shows he ha dwa e- o-sample con amina ion
ans e calcula ed using pos -cleaning ha dwa e back-
g ound da a (Table 4), and by assuming an a e age d illed
sedimen (60 g) ge ing in o con ac wi h a 50 cm segmen
o he d ill s ing (616 cm
2
), 100% o he scoop (217 cm
2
),
and unnel (196 cm
2
), and 100% con amina ion ans e
(d ill o scoop o unnel).
6. Discussion
6.1. E ec i eness o ARADS me hods
We de eloped in eg a ed CCSI p ac ices o ARADS o
p e en and mi iga e con amina ion o he SHTS and he
acqui ed samples and lea n abou po en ial con amina ion
pa hways ac oss he ope a ing en i onmen s. Table 10 sum-
ma izes he mi iga ion asks, key indings, and hei appli-
cabili y o ield condi ions. The ARADS p ac ices a e
simila o hose es ablished o mi iga e con amina ion du ing
a landed mission o Ma s (Sp y e al., 2021).
A e y close ela ionship exis s be ween mic obial educ-
ion and o ganics emo al o p o ec science. Backg ound
con aminan s in spacec a ha dwa e pose wo isks o a li e
de ec ion mission d illing in o he ma ian ice-cemen g ound
Table 10. Desc ip ion and Field Applica ion o Ro e As obiology D illing S udies Mi iga ion P ac ices
(I ems #1–6) and C i ical Resul s
PPE =pe sonal p o ec i e equipmen .
ARADS CONTAMINATION CONTROL STRATEGY IMPLEMENTATION 1323

like Iceb eake (Heldmann e al., 2013; McKay e al., 2013)
and he Ma s Li e Explo e (NAP, 2021).
Fi s , o wa d con amina ion om e es ial mic obes
su i ing spacec a ’s ha dwa e s e iliza ion is he p ima y
conce n (e.g., NRC, 2006; NAS, 2018, 2019, 2020). Spo es
can emain long- e m iable in space, e en wi hou me a-
bolic ac i i y a he low empe a u es o Ma s’ su ace (e.g.,
Ho neck e al., 2010, 2012; Hallswo h, 2021), be shed on
he su ace du ing landing (Ma shall and Mancinelli, 2011),
and en e he unde g ound, becoming me abolically ac i e
unde inc eased wa e ac i i y (Rummel e al., 2014; O osei
e al., 2018); his will inc ease he o wa d con amina ion
isk should mo e habi able condi ions exis unde g ound
o nea he su ace Ca ego y IV biobu den le el limi s a e
£30 +(2 ·10
5
) spo es om he landed spacec a ’s su -
ace, ma ed, and encapsula ed ha dy biobu den su i ing
decon amina ion.
Second, whole mic obes o cellula agmen s could elease
mission- ele an analy es such as amino acids—o any o he
a ge bioma ke along he ‘‘ladde o li e de ec ion’’ (Wes all
e al., 2015; Ne eu e al., 2018)—in o he analy ical sui e. Fo
ins ance, amino acids a e a p ima y a ge o he Iceb eake
li e analyze s and o he li e de ec ion missions.
Bacillus spo es con ain ee (l-glu ama e and l-a ginine)
and bound amino acids, ha is, mucopep ide and p o ein
(Nelson and Ko nbe g, 1970; and e e ences he ein) e-
leasable on agmen a ion. I disloca ed and ans e ed o
analysis, only one B. sub ilis spo e in he space ha dwa e
could elease mo e han 0.24 pg (o 10
-12
g) o al AA.
6.1.1. False nega i es in cleaning e i ica ion. We dem-
ons a ed ha alse nega i e de ec ion o con aminan s
unde mines mic obial educ ion and e i ica ion, esul ing in
mo e conce n o ARADS han alse posi i es, ha is, he
le el o eco e y o swabbed ATP decoupled om he ac-
ual biobu den backg ound. False nega i es can o igina e
om many con ounding ac o s. They a e:
(1) The ha dwa e’s su ace cha ac e is ics (discussed in
Sec ion 6.1.3).
(2) The amoun , dis ibu ion, and complexi y o ha d-
wa e biobu den (bio ilms-o ganics laye ing) and
mine al coa ings (dus o oxidized). When non isible
o ganics and he mine al ilm we e p esen on he d ill,
he ATP swab assay yielded alse nega i es o wo
plausible easons. (i) Mud pa icles and oily esidues
obscu ed he L-L ATP eac ion, and (ii) molecula o -
ganic laye s encasing he mic obes p o ec ed hem
agains de ec ion (Fig. 6C, D) and mic obial educ ion.
The exposed ac i e biobu den could be de ec ed and
educed only a e ace one emo es ino ganic and o -
ganic polyme s (oils, g ease, lipids) be o e mic obial
educ ion. This p oblem did no conce n scoop and
unnels, whose biobu den dec eased linea ly a e ea -
men s (Fig. 6G, H). Unlike he d ill, no complex o ganic
bio( ilms) build-up occu ed du ing he long- e m s o -
age o scoop and unnels.
The cleaning es esul s we e consis en wi h he
common wisdom ha he cleane a su ace can ge ,
he mo e success ul mic obial educ ion and e i i-
ca ion will be (e.g., Mesza os e al., 2005).
Gi en wha has been s a ed ea lie , we sugges
pe o ming mic obial educ ion and e i ica ion on an
ace one-cleaned d ill. Disin ec ion- esis an bio ilm-
o ganics laye ing can be a common issue because he
d ill sys em in gea s, bea ings, and sliding su aces o
he d ill ube connec ions makes ou ine use o s an-
da d lub ican s, o example, 3-in-1 SAE 20 mo o oil
con aining pe oleum, Lub iMa ic Whi e Li hium
G ease, and WD-40 (Mille e al., 2008).
(3) Cleaning agen s’ esidues on ha dwa e. We i s mi i-
ga ed he alse nega i e issue (a de ec ion one) by
conduc ing e i ica ion assays on sol en - insed
ha dwa e. Rinsing wi h wa e emo es in e e ing
cleaning and disin ec ing agen s and add esses one
limi a ion posed o he ATP assay in clean ooms (see
e iew by Shama and Malik, 2013).
(4) Luminome y quenching e ec s by ch omogenic pig-
men s in samples dec ease he ligh emission. Nano-
phase clays, FeOx, and sal s can in e e e wi h L-L
eac ions and chemiluminescence pho ode ec o s.
(5) En i onmen al ield condi ions. High diu nal em-
pe a u es (‡25–40C) and di ec sunligh can igge
e y high RLU alues gene a ing alse posi i es
(Bonacco si and S oke , 2008). The use o chemilu-
minescence pho ode ec o s o e pho omul iplie
FIG. 12. ATP bioma ke s ( o al,
ee, and mic obial) in co ed e sus
su ace sedimen s and de i ed alues
o mic obial biomass as cells/g sedi-
men . No e he negligible con ibu ion
o he po en ial con amina ion ans e
om ha dwa e (To al ATP, sho es
ba s) o d ill samples. 23.7 –2.1
amoles/g o DP and 63.4 –28 amoles/g
(6-Sol Bkg ) Bkg : pos -cleaning ha d-
wa e backg ound. DP, deep cleaning.
1324 BONACCORSI ET AL.
ubes can mi iga e he la e issue. Fu he , wind-
gene a ed iboelec ic e ec s could p oduce unusu-
ally high RLU alues e en a nigh , as we expe i-
enced du ing ield measu emen s. We included
posi i e and nega i e con ols o add ess he assay
pe o mance.
6.1.2. E ec i eness o mic obial educ ion and e i ica-
ion. Coun less cleaning p o ocols exis o labo a o y,
clean oom applica ions o asep ic d illing o icy ma e ials (e.g.,
Ch is ne e al., 2005; Kuhn e al., 2014; Goo dial e al., 2017;
Kayani e al., 2018; Coelho e al., 2022). Ye , only a ew ha e
been designed o complex as obiology li e-de ec ion ech-
nology ials. Mille e al. (2008) applied a mul i-s ep p o ocol
o asep ic d illing (Milli-Q wa e , 10% Lysol aqueous solu ion,
70% E -OH, lame s e iliza ion) (Table 3) and immunoassay-
based deep biosphe e cha ac e iza ion in he Rio Tin o egion
(Pa o e al., 2008), consis en ly wi h o he mul i- eagen
cleaning p o ocol ce i ied by immunoassay-based Limulus
Amebocy e Lysa e (LAL) and GC-MS swab analyses (Eigen-
b ode e al., 2009).
ARADS mul i- eagen p o ocols combined immunoassay
and ATP luminome y echniques o de ec a b oade ange
o iable and non iable biological con aminan s. O e all,
he 6-h mic obial educ ion p o ocol enabled a 4-log d op
o ATP backg ound om unmi iga ed o asep ic condi ions
(1–2 amoles/cm
2
e e ence backg ound wi hin he asep ic
h eshold o £3–4 bac e ial cells/cm
2
); see Sec ion 2.
Du ing he six Sols, asep ic d illing was possible by
ex ending he p o ocol ime (a e Sol 2) ac oss he ime-
cons ained d illing simula ion low (Fig. 7), and consis-
en ly wi h o he mul i- eagen cleaning p o ocols (Mille
e al., 2008; Eigenb ode e al., 2009).
Ac oss he 6-day simula ion, we es ed NaClO-ac i a ed
H
2
O
2
o mo e e ec i e mic obial educ ion. A e squi ing
3% H
2
O
2
on a d ill ip sp ayed wi h NaClO, he esul ing
single oxygen (
1
O
2
) s e ilan agen —decon amina ed he
ha dwa e 30 imes as e han bleach alone o le els
achie able du ing he ex ensi e 6-h deep cleaning in he lab
(Table 4).
The ac i a ion o H
2
O
2
by bleach is an easy and ela i ely
sa e, ield-e ec i e p ocedu e o educing biobu den a e
only a 2-min exposu e, compa ed wi h he long exposu e
equi ed by concen a ed bleach alone (10–20¢), 10% bleach
(3 h), 70% IPA (1 h), o 3% H
2
O
2
alone (>3 h).
The edox eac ion be ween he oxidizing hypochlo i e ion
(OCl
-
) and he educing H
2
O
2
agen o ms wa e (H
2
O), chlo-
ine ion (Cl
-
), and
1
O
2
(Held e al., 1978; G ee , 2006) such as:
H2O2(aq)þNaCIO (aq)!1O2(g)þNaCI (aq)þH2O(aq)
The disin ec ion ac ion was due o he exci ed s a e o
1
O
2
, which con ains mo e ene gy han g ound oxygen (Held
e al., 1978; Wi z, 2009).
1
O
2
un olds p o eins (e.g., Del
Maes o e al., 1980; Da ies, 2003) and kills bac e ial and
euka yo ic cells (Dahl e al., 1987; Djimeli e al., 2014;
Taewan e al., 2019) wi h mic oo ganism inac i a ion p o-
po ional o
1
O
2
exposu e.
Low- empe a u e chemical s e ilan s such as 35% H
2
O
2
Gas Plasma umiga ion, E hylene Oxide (E O), and Ozone
(O
3
) a e all absolu e biocides commonly used in clean
ooms (e.g., Go don e al., 2012). Howe e , hese ech-
niques a e inapplicable in he ield because hey a e highly
oxic and equi e complex equipmen (e.g., con ainmen
chambe s, deep acuum d aw zapping, gas moni o ing
sys ems) and se e al hou s-exposu e (Mesza os e al., 2005;
Go don e al., 2012).
6.1.2.1. Cleanliness e i ica ion and con amina ion
knowledge. We used wo me hods o cleanliness e i ica-
ion, he ATP Assay, and he SOLID ins umen mic oa ay
immunoassay echnology, as a no el app oach o con am-
ina ion knowledge o he ha dwa e backg ound. We could
no use he cul u e-based NSA, which enume a es (pla e
coun ing) d y hea -su i ing ae obes endospo es a e a 72 h
g ow h beyond he ARADS eal- ime imeline.
In addi ion, he NSA o e looks non-cul i able cells,
unde es ima ing he o al mic obial bu den, spo e- o ming
s ic anae obes (Clos idium), and ex emophilic a chaea
mic oo ganisms o PP conce n (e.g., Moissl-Eichinge e al.,
2015). Fo hese easons, Be na dini and Venka eswa an
(2016) co-analyzed ATP and NSA da a om he MSL
spacec a .
They p oposed he ATP assay (pe NASA-HDBK-6022,
2010) as he benchma k h eshold o spacec a su ace
cleanliness, ha is, 2.3 ·10
-11
mmol ATP/25 cm
2
(o <0.9
mole/cm
2
). Se e al me hods exis o de ec and cha ac e ize
ha dwa e con aminan s ma ix-assis ed lase deso p ion/io-
niza ion- ime o ligh , bu complex se ups limi hei use in
he ield. O he eal- ime ools designed o ield de ec ion,
such as Minion Ox o d Nanopo e o DNA and RNA se-
quencing (10 min) o he LAL assay o G am-nega i e
mic oo ganisms, ha e been used (pos -mission) in ield i-
als (Eigenb ode e al., 2009; Cockell e al., 2018).
Howe e , he LAL assay does no de ec only iable
mic obes and equi es mo e ex ended sample p ocessing
and analysis ime han he ATP Luminome y assay, ha is,
15–60¢ e sus 15 s -5 min, espec i ely. Thei applica ion
is beyond he scope o he quick disin ec ion alida ion
equi emen ac oss he six Sols.
In si u eal- ime and ul asensi i e ATP assay can de ec
0.1 moles o ATP on ha dwa e su ace as an independen
molecula p oxy o ac i e biobu den/mic obial biomass,
iable spo es, o in ac ecen ly dead cells. Fu he , he ATP
syn hase FI signal was consis en wi h ATP’s pos -cleaning
ha dwa e backg ound le els. This independen e i ica ion
co obo a es he e ec i eness o in-sim cleaning and
mic obial educ ion.
Fu he , he luo escence signal agains he p o ein bio-
ma ke s was highe han ha agains he whole cells. The
wo signals decoupled, likely because mo e agmen ed
cellula ma e ials e sus whole cells we e p esen in he
wind-ca ied dus (aeolian pa icles subjec o pho o-
oxida ion) and subsu ace sedimen .
SOLID immunog ams o whole cells, p o eins, and o he
cellula compounds p o ided an e ec i e e i ica ion ool
o sel -check agains alse posi i es con amina ing science
measu emen s. One limi a ion is ha he immunoassay does
no di e en ia e be ween li ing o dead cells (a o wa d con-
amina ion isk), bu ATP assay co-analysis add esses his gap.
Finally, GC-MS and high-pe o mance liquid ch oma o-
g aphy iden i y backg ound con aminan s in space ha dwa e
(con amina ion knowledge) wi h subs an ial ime elapsed
be ween sampling and analysis. Fo he ARADS ha dwa e
ARADS CONTAMINATION CONTROL STRATEGY IMPLEMENTATION 1325
con amina ion knowledge, we used SOLID immunog ams;
such an app oach could educe ime and cos s o he si-
mul aneous assessmen o o wa ding and science con am-
ina ion.
6.1.3. Role o ha dwa e su ace ea u es o geome y.
The space ha dwa e design, geome y, su ace opog aphy,
po osi y, and oughness can a ec con amina ion con ol
e ec i eness. Fo he same eason, hese ea u es can
de e mine biobu den e en ion, dislodgemen , ans e , o
dispe sal o he en i onmen . Speci ically, he s ainless-s eel
a m-moun ed Phoenix-like scoop in e io was di icul o
access due o i s 90edges be ween he in e io su aces.
The su ace o he ex e nal scoop blade was mo e accessible
bu oughe han he scoop in e io . As a esul , he scoop’s
in e io and edge exhibi ed a pos -cleaning h eshold back-
g ound highe han he unnels.
Simila ly, he TRIDENT d ill bi ’s inishing was h ee-
dimensional (3D) p in ed and e ched o augmen he
eco e y o d illed ines. This d ill bi su ace was oughe
han he d ill s ing (Fig. 2), esul ing in high biobu den
backg ound (Fig. 6G, H and Tables 4 and 5). Howe e , an
augmen ed cleaning p o ocol educed he backg ound.
Mi iga ing solu ions could include cu en good manu ac-
u ing p ac ices such as ‘‘co ing’’ and polished su ace in-
ishing. Co ing minimizes geome y- ela ed con amina ion by
eplacing 90and 3D co ne s wi h cu ed su aces and im-
p o es s e iliza ion o clean ooms and hospi al loo s. Cu ed
co ne s inside he scoop su aces (inne walls) would p e en
he accumula ion o biological con aminan s o d illed ma e-
ial (conduci e o c oss-con amina ion) and acili a e he
pa hway o cleaning and swabbing.
An addi ional implemen a ion could use s a e-o - he-a
mi o -like polished su aces o he d ill and SHTS. Elec-
opolishing ensu es ul a cleaning o embedded con ami-
nan s in Ti anium alloys o issue enginee ing and su gical
implan s, which mus be s e ile be o e use. In addi ion,
elec opolished mic oscopically ea u eless su aces educe
mic obial adhesion and su ace con amina ion (Bagno and
Bello, 2004; Oshida, 2007; Shimaku a, 2007; Tajima e al.,
2008).
6.2. Con amina ion pa hways
We p opose a concep ual model (Fig. 13) ha includes
he signi ican con amina ion pa hways highligh ed by da a
analysis and es s on ARADS ma e ials (ha dwa e, sedi-
men s, ai sample s, e c.). In e es ingly, he s udy o
ai bo ne ma e ials deli e ed wo su p ising esul s: a highe -
han-p edic ed en i onmen al con amina ion (pa hway 8–9
o 3–4); and a lowe - han-expec ed an h opogenic one
(pa hway 1–2 o 3). The eme ging pic u e is ascina ingly
complex due o he in insic na u e o he A acama Dese .
6.2.1. En i onmen al sou ces o ai bo ne con aminan s.
Wind- anspo ed mine al dus can e icien ly ca y and
dispe se iable mic obial li e ac oss he A acama (Pa o
e al., 2011; and e e ences he ein). Whe he his biomass
cons i u es an an h opogenic (human biome) o na u al
(en i onmen al) con aminan , a c oss-con aminan , o a
‘‘na i e’’ elemen is no always clea o us o unde s ood
(Azua-Bus os e al., 2019, 2022).
Ou es s e ealed ha ai bo ne biomass ep esen ed an
unusually high sou ce o po en ial con amina ion, unex-
pec ed o he hype a id co e o he A acama Dese .
The high-in ensi y signal (>6000)— aised agains aqua ic
mic o lo a (Anabaena o Nos oc sp. cyanobac e ia) in ai
sample s—poin s o a p ima y sou ce o ai bo ne bio-
mass consis en wi h ATP da a om he pos - looded Playa
(10
6
cells/g sedimen , ATP cell equi alen ) and he ai bo ne
FIG. 13. Concep ual model illus a ing
sou ce- o-sink con amina ion. Figu e no
o scale. Semi-quan i a i e cell es ima es
[cells/g, cells/cm
2
,andcells/(cm$day
1
)]
and numbe s nea each pa hway. Base-
camp [1] and an h opogenic con amina-
ion[2]indus Coll.1[3]andColl.2[4].
The hin dashed a ows indica e Ro e [5]
and c oss-con amina ion pa hways:
(a) d ill- o-subsu ace [6] (a o wa d
con amina ion analog); (a–d) d ill-
o-sample ans e and analysis: (a) d ill,
(b) scoop, (c) unnel, (d) li e de ec ion
ins umen . Blue solid-lined a ows wi h
‘‘X’’ indica e unlikely con amina ion
pa hways, o example, desicca ed pos -
lood mic obial ma [8] sou cing aqua ic
cyanobac e ia [9] o sampling ha dwa e
(a–c) o subsu ace sedimen a y laye s [6]
and no looded playa su ace [7] delimi-
ed by he Playa sho eline (black dashed
lines). La ge g een a ows indica e un-
expec ed ai bo ne biology, o example,
Anabaena sp. [9] in collec o s [3, 4]. Blue
and ed pa allel dashed a ows ep esen
he e ogeneous (mo e dis al?) sou ces o
ai bo ne biomass [10] o he collec o s o
sampling ha dwa e. Coll., collec o .
1326 BONACCORSI ET AL.
biomass—5–22 amoles/(cm
2
$day) ATP, ac oss he 6-Sol
mission (Fig. 12).
This sugges s an unexpec edly high iable biomass la e al
ans e om an unusual upwind sou ce, ha is, desicca ed
ponds wi hin he nea by Playa a ew ens o me e s away
om he d ill si e and 300–400 m om Basecamp Collec-
o 1. Such an ai bo ne sedimen a y biomass ep esen s a
po en ial sou ce o high con aminan (also e y close o one
o he d ill si es) ha could ha e en e ed obo ic sampling
and analysis i no mi iga ed.
Epheme al wa e -induced mic o lo a popula ion g ow h is
uncommon and ep esen s a no el phenomenon o he
hype -a id co e o he A acama Dese due o he ecen
onse (in 2015) o clima ic change b inging yea ly s o ms
and loods o he A acama egion (Fe na
´ndez-Ma ı
´nez
e al., 2019; K. Wa en-Rhodes 2022, unpublished da a).
Howe e , bu s s o mic o lo a g ow h can occu a e ex-
eme ain e en s in a id ho dese en i onmen s wo ldwide
(e.g., Ci e
´se al., 2017; and e e ences he ein; Bonacco si
and McKay, 2022).
The weake -in ensi y signals (£1000) wi h LDChip poin
o seconda y low-in ensi y sou ces o he han he Playa. Fo
ins ance, anspo om he A acama soil mic oen i on-
men s con aining s ong acids (Quinn e al., 2005) could
explain he de ec ion o bac e ia om acidic habi a s (Pa o
e al., 2011; and e e ences he ein). Simila ly, he de ec ed
a chaea Me hanobac e ium could ha e been wind-ca ied
om he same sou ce o he aqua ic cyanobac e ia, he pos -
looded Playa.
Me hanogenesis can occu a he wa e -sedimen in e -
ace, in anoxic mic opo es o ine-g ained clay- ich sedi-
men s du ing loods. Gammap o eobac e ia (Pseudomonas)
and hei associa ed bioma ke a e ound in ex eme desic-
ca ed en i onmen s (Pa o e al., 2011), including A acama
playas and allu ial an samples (Fe na
´ndez-Ma ı
´nez
e al., 2019), subsu ace hype saline habi a s (e.g., Sala
G ande). The anae obic Shewanella inhabi s hea y me al-
con amina ed (i on, lead, and u anium) en i onmen s
(mine ac i i y). All hese en i onmen s such as he GP
Si e (Fig. 4D, E) exis in closed o dis al p oximi y o he
ARADs’ d ill si es and can p o ide biological con ami-
nan s o conce n depending on hei ela i e abundance and
iabili y.
The hype -a id A acama’s su ace soil is mos ly ex e-
mely low in iable mic obes and associa ed biomolecula
compounds (e.g., Came on, 1969; Na a o-Gonzalez e al.,
2003; Skelley e al., 2005; Wa en-Rhodes e al., 2006;
Ca
´ce es e al., 2007; Cockell e al., 2008) wi h biomass
inc easing wi h some dep h (e.g., Fe na
´ndez-Ma ı
´nez e al.,
2019). A ew excep ions can include ex ensi e su ace
sou ce a eas such as sala s, mic o lo a-colonized hali e
ields (e.g., Da ila e al., 2010; Pa o e al., 2011; Wie zchos
e al., 2006, 2011; Da ila and Schulze-Makuch, 2016),
g oundwa e - ed oases wi h spa se ascula plan s (Fig. 4D, E),
o mic o lo a blooms in esponse o spo adic and ansien
ain e en s (e.g., Schulze-Makuch e al., 2018; U i skiy
e al., 2018; 2019).
Ai bo ne dus om he sou ces ci ed ea lie migh ca y
con aminan s o d illed samples o en e he analy ical
ins umen , hus con using he scien i ic esul s. A second-
a y bu no less impac ul con aminan s a e ‘‘ esh’’ and
deg aded ossilized biomolecules, o example, ATP,
Phospholipid a y acids (PLFA), DNA, lipopolysaccha ides
(LPS), which he hype -a id su ace soil o he A acama
Dese can p ese e well (Les e e al., 2007; Bonacco si
e al., 2010; Powe s e al., 2018; Wilhelm e al., 2018).
6.2.2. An h opogenic ai bo ne con amina ion. We mon-
i o ed he lux o ai bo ne con aminan s du ing he in-sim
ope a ions o e alua e he an h opogenic con amina ion
isk (expec ed high) o he Playa en i onmen and ARADS
science. The biomass lux a he an h opogenic-domina ed
basecamp a ea was highe [22.1 amoles/(cm
2
$day) ATP]
han ha moni o ed by Collec o 1, 250 m away om
basecamp [5.2 amoles/(cm
2
$day) ATP; non-an h opogenic
sou ces].
This lux o ATP-based biomass was highe han he
a e age ATP backg ound o he (pos -cleaning) SHTS
ha dwa e, ha is, 0.9 –1.1 o 3.6 –4.5 amoles/cm
2
(Table 5),
implying ha we e ec i ely mi iga ed he deli e y o ai -
bo ne con aminan s o he ha dwa e. Howe e , a Basecamp,
LDChip200 de ec ed mic oo ganisms na u ally occu ing in
he A acama Dese (ai bo ne om p oximal and dis al
po en ial sou ce a eas).
Only a ew axa de ec ed wi h a weak signal (FI <1000)
(B. sub ilis spo es; S ep omyces spp., spo e, and mycelium)
migh be ela ed o he human mic obiome. Howe e ,
B. sub ilis and S ep omyces spp. a e usual bac e ia om
soil (also common in he A acama). B.sub ilis spo es can
su i e o up o 15 mon hs in he hype -a id co e o he
A acama Dese wi h 15% su i al (Dose e al., 2001).
S ep omyces is a ilamen ous ac inobac e ium na u ally
p esen in he soil. This alkaliphilic and he mophilic genus
is ound in a id, hype saline, o hea y me als- ich ex eme
habi a s (Kamp e e al., 2014), including he hype -a id
A acama (San hanam e al., 2012; Good ellow e al., 2017).
S ep omyces can also be ound in a heal hy human
mic obiome (a a e occu ence), whe e cul u e-based ech-
niques ha e o e looked i s p esence due o low g ow h, bu
ecen molecula -based s udies highligh ed i s p esence in
he gas o-in es inal (Bolou ian and Moj ahedi, 2018) and
espi a o y ac s (Huang e al., 2015; He b ı
´ke al., 2020)
and in he heal hy skin (Gallo and Hoope , 2012). In pa -
icula , skin lakes a e ich in soma ic cells (5 ·10
8
cells
shed e e y day o 2 ·10
7
cells e e y hou (Weschle , 1978))
and associa ed mic obes.
Fo ins ance, he hand mic obio a a ies *4·10
4
o
4.6 ·10
6
CFU/cm
2
(Leyden e al., 1987; Message e al.,
2004) and 1 ·10
7
cul u able bac e ia pe cm
2
(F ed icks,
2001); ye hese cul u e-based es ima ions a e likely unde -
es ima es (Edmonds-Wilson e al., 2015; and e e ences
he ein).
On a cau iona y no e, Bacillus and S ep omyces axa
could come om he en i onmen a he han human biome
con aminan s. Howe e , we canno disca d his hypo hesis
ei he .
An explana ion o he weakness o he ai bo ne an h o-
pogenic signal could be ine icien sampling (unde -
sampling) o ai bo ne pa icles, ha is, he swab was
e ically placed o collec dus pa icles di ec ly om he
ai . Ano he possibili y is ha he an h opogenic con ami-
nan s (skin, clo hing, and d ople s) shed by he human
c ew du ing ope a ions we e non-deposi ed, emo ed a e
deposi ion, o dispe sed by ai low and wind. Fu he , he
ARADS CONTAMINATION CONTROL STRATEGY IMPLEMENTATION 1327
signs o li e de ec o -li e de ec o chip du ing a Ma s d illing
simula ion campaign in he hype a id co e o he A acama
Dese . As obiology 2023;23(12); doi: 10.1089/as .2021
.0174
NAP. Plane a y Science and As obiology Decadal Su ey.
Ma s Li e Explo e Mission Concep S udy Repo , 2021.
NASA-HDBK-6022. Handbook o he Mic obial Examina ion
o Space Ha dwa e. Na ional Ae onau ics and Space Admin-
is a ion: Washing on, DC; 2010.
Na ional Ae onau ics and Space Adminis a ion (NASA). Pla-
ne a y P o ec ion P o isions o Robo ic Ex a e es ial
Missions. NPR 8020.12D, Na ional Ae onau ics and Space
Adminis a ion: Washing on, DC; 2011.
Na ional Academies o Sciences, Enginee ing, and Medicine
(NAS); Space S udies Boa d; Commi ee on he Re iew o
Plane a y P o ec ion Policy De elopmen P ocesses. Re iew
and Assessmen o Plane a y P o ec ion Policy De elopmen
P ocesses. The Na ional Academies P ess: Washing on, DC;
2018; doi: 10.17226/25172
Na ional Academies o Sciences, Enginee ing, and Medicine
(NAS); Space S udies Boa d; Commi ee o Re iew he
Repo o he NASA Plane a y P o ec ion Independen
Re iew Boa d. NASA Plane a y P o ec ion Independen
Re iew Boa d (PPIRB) Repo o NASA/SMD. 2019.
Na ional Academies o Sciences, Enginee ing, and Medicine
(NAS); Space S udies Boa d; Commi ee o Re iew he
Repo o he NASA Plane a y P o ec ion Independen
Re iew Boa d. Assessmen o he Repo o NASA’s Plane a y
P o ec ion Independen Re iew Boa d. The Na ional Acade-
mies P ess: Washing on, DC; 2020; doi: 10.17226/25773
Na ional Resea ch Council (NRC). P e en ing he Fo wa d
Con amina ion o Ma s. The Na ional Academies P ess:
Washing on, DC; 2006. doi: 10.17226/11381
Na a o-Gonzalez R, Rainey FA, Molina P, e al. Ma s-like
soils in he A acama Dese , Chile, and he d y limi o
mic obial li e. Science 2003;302:1018–1021.
Nelson DL, Ko nbe g A. Biochemical s udies o bac e ial
spo ula ion and ge mina ion. 18. F ee amino acids in spo es.
J Biol Chem 1970;245(5):1128–1136.
Nesce ecka A, Juhna T, Hammes F. Beha io and s abili y o
adenosine iphospha e (ATP) du ing chlo ine disin ec ion.
Wa e Res 2016;101:490–497; doi: 10.1016/j.wa es.2016.05
.087
Ne eu M, Hays LE, Voy ek MA, e al. The ladde o li e
de ec ion. As obiology 2018;18(11):1375–1402; doi: 10
.1089/as .2017.1773
Okanojo M, Miyashi a N, Tazaki A, e al. A omol-le el ATP
bioluminome e o de ec ing single bac e ium. Luminescence
2017;32:751–756; doi: 10.1002/bio.3246
O osei R, Lau o SE, Pe inelli E, e al. Rada e idence o
subglacial liquid wa e on Ma s. Science 2018;361(6401):
490–493.
Oshida Y. Su ace science and echnology—Ti anium den al im-
plan sys ems. J Soc Ti anium Alloys Den is 2007;5:52–53.
Pandey S, Cla ke J, Nema P, e al. Ladakh: Di e se, high-
al i ude ex eme en i onmen s o o -Ea h analogue and
as obiology. In J As obiol 2019;19(1):1–21; doi: 10.1017/
S1473550419000119
Panousi MN, Williams GJ, Gi dles one S, e al. E alua ion o
alcohol wipes used du ing asep ic manu ac u ing. Soc Appl
Mic obiol Le Appl Mic obiol 2009;48:648–651.
Pa nell J, Cullen D, Sims M, e al. Sea ching o li e on Ma s:
Selec ion o molecula a ge s o ESA’s Au o a ExoMa s
mission. As obiology 2007;7:578–604.
Pa o V, Fe na
´ndez-Cal o P, Rod ı
´guez Man edi JA, e al.
SOLID2: An an ibody a ay-based li e de ec o ins umen in
a Ma s D illing Simula ion Expe imen (MARTE). As o-
biology 2008;8(5):987–999.
Pa o V, de Diego-Cas illa G, Mo eno-Paz M, e al. A mic obial
oasis in he hype saline A acama subsu ace disco e ed by a
li e de ec o chip: Implica ions o he sea ch o li e on Ma s.
As obiology 2011;11(10):969–996.
Pa o V, Puen e-Sa
´nchez F, Cab ol NA, e al. Mic obiology and
ni ogen cycle in he ben hic sedimen s o a glacial oligo ophic
deep Andean lake as analog o ancien ma ian lake-beds. F on
Mic obiol 2019:10:929; doi: 10.3389/ micb.2019.00929
Pa el A, Malino ska L, Saha S, e al. ATP as a biological
hyd o ope. Science 2017;356:753–756.
Powe s LS, Smi h HD, Kilungo AP, e al. In si u eal- ime
quan i ica ion o mic obial communi ies: Applica ions o cold
and d y olcanic habi a s. Glob Ecol Conse 2018;16:e00458.
Quinn R, Zen AP, G un hane F, e al. De ec ion and cha ac-
e iza ion o oxidizing acids in he A acama Dese using he
Ma s Oxida ion Ins umen . Plane Space Sci 2005;53(13):
1376–1388; doi: 10.1016/j.pss.2005.07.004
Ri as LA, Ga cı
´a-Villadangos M, Mo eno-Paz M, e al. A 200-
an ibody mic oa ay biochip o en i onmen al moni o ing:
Sea ching o uni e sal mic obial bioma ke s h ough immu-
nop o iling. Anal Chem 2008;80;7970–7979.
Rummel JD, Bea y DW, Jones MA, e al. A new analysis o
Ma s ‘‘special egions’’: Findings o he second MEPAG
special egions science analysis g oup (SR-SAG2). As o-
biology 2014;14:887–968.
Ru ala WA, Webe DJ; he Heal hca e In ec ion Con ol P ac-
ices Ad iso y Commi ee. Guideline o Disin ec ion and
S e iliza ion in Heal hca e Facili ies. 2019; p. 163. A ailable
om: h ps://www.cdc.go /in ec ioncon ol/pd /guidelines/
disin ec ion-guidelines-H.pd [Las accessed: Decembe 11,
2023].
Sanchez-Ga cı
´a L, Fe na
´ndez-Ma ı
´nez MA, Mo eno-Paz M,
e al. Simula ing Ma s d illing mission o sea ching o li e:
G ound- u hing lipids and o he complex mic obial bio-
ma ke s in he i on-sul u ich Rı
´o Tin o analog. As obiology
2020;20(9):1029–1047; doi: 10.1089/as .2019.2101
San hanam R, Oko o CK, Rong X, e al. S ep omyces a aca-
mensis sp. no ., isola ed om an ex eme hype -a id soil o
he A acama Dese , Chile. In J Sys E ol Mic obiol 2012;62:
2680.
Schulze-Makuch D, Wagne D, Kouna es SP, e al. T ansi o y
mic obial habi a in he hype a id A acama Dese . P oc Na l
Acad Sci U S A 2018;115(11):2670–2675; doi: 10.1073/
pnas.1714341115
U i skiy G, Ge sin S, Munn, A, e al. Response o ex emophile
mic obiome o a a e ain all e eals a wo-s ep adap a ion
mechanism. bioRxi 442525, 2018; doi: 10.1101/442525
Shama G, Malik DJ. The uses and abuses o apid
bioluminescence-based ATP assays. In J Hyg En i on
Heal h 2013;216:115–125.
Shimaku a M. E icien polishing me hod o i anium. J Soc
Ti anium Alloys Den is 2007;5:32–33.
Si agusa SR, Do sa WJ, Cu e CN, e al. Use o a newly
de eloped apid mic obial ATP bioluminescence assay o
de ec mic obial con amina ion on poul y ca casses. J Bio-
luminesc Chemiluminesc 1996;11(6):297–301.
Skelley AM, Sche e JR, Aube y AD, e al. De elopmen and
e alua ion o a mic ode ice o amino acid bioma ke
de ec ion and analysis on Ma s. P oc Na l Acad Sci U S A
2005;102:1041–1046.
1334 BONACCORSI ET AL.

Smi h SA, Bena dini JN, Ande l D, e al. Iden i ica ion and
cha ac e iza ion o ea ly mission phase mic oo ganisms
esiding on he Ma s Science Labo a o y and assessmen o
hei po en ial o su i e Ma s-like condi ions. As obiology
2017;17(3):253–265; doi: 10.1089/as .2015.1417
So en GA, Snyde CW. Fi s Viking mission o Ma s. Science
1976;193:759–766.
Sp y JA, Siegel B, Kminek G, e al. Plane a y p o ec ion
knowledge gaps and enabling science o human Ma s mis-
sions. Bull Am As on Soc 2021;53(4):205.
S anley PE. Ex ac ion o adenosine iphospha e om mic o-
bial and soma ic cells. Me hods Enzymol 1986;133:14–22.
S oke CR, Cannon HN, Dunagan SE, e al. The 2005
MARTE Robo ic D illing Expe imen in Rio Tin o Spain:
Objec i es, app oach, and esul s o a simula ed mission o
sea ch o li e in he Ma ian subsu ace. As obiology
2008;8(5):921–945.
S oke CR, Glass BJ, Glass, S ucky TR, e al. A mission si-
mula ing he sea ch o li e on Ma s wi h au oma ed d illing,
sample handling, and li e de ec ion ins umen s pe o med in
he hype a id co e o he A acama Dese , Chile. As obiology
2023;23(12):1284–1302; doi: 10.1089/as .2022-0055
Taewan K, Hyung-eun K, Jiyoon C, e al. Inac i a ion o
Esche ichia coli and MS2 coliphage ia single oxygen
gene a ed by homogeneous pho osensi iza ion. Ko ean J
Chem Eng 2019;36(11):1785–1790; doi: 10.1007/s11814-
019-0353-4
Tajima K, Hi onaka M, Chen K, e al. Elec opolishing o CP
i anium and i s alloys in an alcoholic solu ion-based elec-
oly e. Den Ma e J 2008;27(2):258–265.
U i skiy G, Ge sin S, Munn A, e al. Halophilic mic obial
communi y composi ional shi a e a a e ain all in he
A acama Dese . ISME J 2019;13(11):2737–2749; doi: 10
.1038/s41396-019-0468-y
Venka eswa an K, Ha o i N Duc MTL, e al. ATP as a bio-
ma ke o iable mic oo ganisms in clean- oom acili ies.
J Mic obiol Me hods 2003;52(3):367–377.
Wa en-Rhodes KA, Rhodes KL, Poin ing SB, e al. Hypoli hic
cyanobac e ia, d y limi o pho osyn hesis, and mic obial
ecology in he hype -a id A acama Dese . Mic ob Ecol 2006;
52:389–398.
Wa en-Rhodes KA, Lee KC, A che SDJ, e al. Subsu ace
mic obial habi a s in an ex eme dese Ma s-analog en i-
onmen . F on Mic obiol 2019;10:69; doi: 10.3389/ micb
.2019.00069
Weschle CJ. Cha ac e iza ion echniques applied o indoo
dus . En i on Sci Technol 1978;12(8):923; doi: 10.1021/
es60144a011
Wes all F, Fouche F, Bos N, e al. Biosigna u es on Ma s:
Wha , whe e, and how? Implica ions o he sea ch o ma -
ian li e. As obiology 2015;15:998–1029.
Wie zchos J, Ascaso C, McKay CP. Endoli hic cyanobac e ia in
hali e ocks om he hype a idco e o he A acama Dese .
As obiology 2006;6:415–422.
Wie zchos J, Cama a B, de los Rios A, e al. Mic obial colo-
niza ion o Ca-sul a e c us s in he hype a id co e o he
A acama Dese : Implica ions o he sea ch o li e on Ma s.
Geobiology 2011;9(1):44–60; doi: 10.1111/j.1472-4669.2010
.00254.x
Wilhelm MB, Da ila AF, Pa en eau MN, e al. Cons ain s on
he me abolic ac i i y o mic oo ganisms in A acama su ace
soils in e ed om e ac o y bioma ke s: Implica ions o
ma ian habi abili y and bioma ke de ec ion. As obiology
2018;10(7):955–966.
Willis P, C eame JS, Mo a M. Implemen a ion o mic ochip
elec opho esis ins umen a ion o u u e space ligh mis-
sions. Anal Bioanal Chem 2015;407:6939–6963.
Willson D, Bonacco si R, Gold R, e al. Con amina ion Man-
agemen o Sample Collec ion De ices and Ocean Analogs
o Li e De ec ion on Icy Wo ld Plume Fly- h ough Missions.
Pape ID. B5.3-64-18. 42nd COSPAR Scien i ic Assembly,
July 14–22, 2018, Pasadena, CA, 2018.
Wi z KPJ. Pho ochemis y o O ganic Compounds: F om Con-
cep s o P ac ice. John Wiley & Sons: Chiches e , UK; 2009.
Zwa k uis E, Be ay G, Wilke T. Rapid es ing o mic obial
su ace con amina ion on eal ca casses using ATP biolu-
minescence. Fleischwi scha 1999;79:101–103.
Add ess co espondence o:
Rosalba Bonacco si
NASA Ames Resea ch Cen e MS. 245-3
Mo e Field, CA 94305
USA
E-mail: [email p o ec ed]
Submi ed 10 Oc obe 2022
Accep ed 2 July 2023
Associa e Edi o : Jack Mus a d
Abb e ia ions Used
1
O
2
¼single oxygen
3D ¼ h ee-dimensional
ARADS ¼A acama Ro e As obiology D illing S udies
ASB ¼ATP syn hase subuni 1
ASF1 ¼ATP syn hase subuni 2
ATLO ¼Assembly, Tes , and Launch Ope a ions
ATP ¼adenosine iphospha e
A g. ¼a e age
CCSI ¼Con amina ion Con ol S a egy and
Implemen a ion
CFU ¼colony o ming uni
COSPAR ¼Commi ee on Space Resea ch
DHMR ¼d y hea o mic obial educ ion
DP ¼deep-cleaning
DW ¼dis illed/deionized wa e
E -OH ¼e hanol
FI ¼ luo escence in ensi y
FSMI ¼Fluo escence Sandwich Mic oa ay
Immunoassay
FTIR ¼Fou ie ans o m in a ed
GC-MS ¼gas ch oma og aph–mass spec ome e
GNCA ¼G am-nega i e bac e ia ances al be a-lac amase
GP ¼G een Pa o
GPBCA ¼gammap o eobac e ial ances al
be a-lac amase
H
2
O
2
¼hyd ogen pe oxide
HD ¼ha dwa e
IPA ¼isop opyl alcohol
ISAD ¼Icy Soil Acquisi ion De ice
LAL ¼Limulus Amebocy e Lysa e
LC/MS ¼liquid ch oma og aphy/mass spec ome y
LITMS ¼Linea Ion T ap Mass Spec ome e
L-L ¼luci e in-luci e ase
LOD ¼limi o de ec ion
LPS ¼lipopolysaccha ides
Me -OH ¼me hanol
ARADS CONTAMINATION CONTROL STRATEGY IMPLEMENTATION 1335
MILA ¼Mic o luidic Li e Analyze
MSL ¼Ma s Science Labo a o y
ND ¼no de ec ion
NSA ¼NASA S anda d assay
PLFA ¼phospholipid a y acids
PP ¼Plane a y P o ec ion
ppb ¼pa s pe billion
PPE ¼pe sonal p o ec i e equipmen
PSTAR ¼Plane a y Science and Technology
Th ough Analog Resea ch
RLU ¼ ela i e luminosi y uni s
SHTS ¼Sample Handling and T ans e Sys em
SOLID ¼Signs o Li e De ec o
STD ¼s anda d de ia ion
SWA ¼swab analysis
TEGA ¼The mal E ol ed Gas Analyze
TRIDENT ¼The Regoli h and Ice D ill o Explo a ion
o New Te ains
UHV ¼ul a high acuum
WCL ¼We Chemis y Labo a o y
1336 BONACCORSI ET AL.