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Composi ion, s uc u e
and obus ness o Lichen guilds
Sal a Du an‑Neb eda
1* & Se gi Val e de
1,2*
Symbiosis is a majo engine o e olu iona y inno a ion unde lying many ex an complex o ganisms.
Lichens a e a pa adigma ic example ha o e s a unique pe spec i e on he ole o symbiosis in
ecological success and e olu iona y di e si ica ion. Lichen s udies ha e p oduced a weal h o
in o ma ion ega ding he impo ance o symbiosis, bu hey equen ly ocus on a ew species,
limi ing ou unde s anding o la ge‑scale phenomena such as guilds. Guilds a e g oupings o lichens
ha assis each o he ’s p oli e a ion and a e in ima ely linked by a sha ed se o pho obion s,
cons i u ing an ex ensi e ne wo k o ela ionships. To cha ac e ize he ne wo k o lichen symbion s,
we used a la ge da a se (
n=206
publica ions) o na u al pho obion ‑mycobion associa ions.
The en i e lichen ne wo k was ound o be modula , bu his o ganiza ion does no di ec ly ma ch
axonomic in o ma ion in he da a se , p omp ing a econside a ion o lichen guild s uc u e
and composi ion. The mul iscale na u e o his ne wo k e eals ha he majo lichen guilds a e
be e ep esen ed as clus e s wi h se e al subs uc u es a he han as monoli hic communi ies.
He e ogeneous guild s uc u e os e s obus ness, wi h keys one species unc ioning as b idges
be ween guilds and whose ex inc ion would endange global s abili y.
Lichens a e symbio ic o ganisms composed o a ungus (mycobion ), one o mo e pho osyn he ic pa ne s
(pho obion s, ypically algae o cyanobac e ia see Fig.1a, b) and o he mic obial species1,2. In he wide eco-
logical con ex , lichens p o ide se e al se ices ha a e essen ial o ecosys em unc ioning: om wea he ing
o ocks inc easing he bioa ailabili y o mine als3 o ca bon and ni ogen ixa ion4. By i ue o he wildly di -
e en me abolisms o pho obion s and mycobion s, licheniza ion p o ides new biological ai s ha can enable
bo h pa ne s o colonize a wide ange o en i onmen s5,6, including ex eme3,7, pollu ed8,9 o an h opogenic
ecosys ems10. In consequence, lichens a e mo e han he sum o hei cons i uen symbion s, emphasizing he
ele ance o non- a e nal o ganismali y as a sou ce o e olu iona y inno a ion11,12.
Symbiosis is a na u al ho spo o di e si y and inno a ion13,14, he e olu iona y po en ial o a lichen symbion
mus ake in o accoun he ex ao dina y epe oi e o ai s gained by acqui ing a new pa ne . Howe e , i is
widely acknowledged ha speci ici y o he mycobion -pho obion associa ion as well as he spa ial dis ibu ion
o hei componen species la gely d i es he o ma ion o new lichen o ganisms6,15–18. Indeed, many symbion s
a e qui e s ingen in he pa ne ships hey o m, which a e o en de e mined by biophysical g adien s19–24 o
subs a e p e e ences18. Selec i e mycobion s do no always use he en i e niche o hei pho obion s and a e
he e o e some imes es ic ed o ce ain clima ic condi ions. I has also been sugges ed ha highly speci ic
mycobion s a e ypically es ic ed o ew en i onmen s and display limi ed ecological ange25,26. Con e sely,
mycobion s capable o colonizing di e en habi a s o en a e less s ingen in hei pa ne ships, in e ac ing wi h
a ious a ailable pho obion s13,17.
Collec i ely, he se o na u al associa ions be ween pho obion s and mycobion s de ines a ne wo k o
symbion s27,28, whe e each in e ac ion co esponds o a singula lichen species (Fig.1c). I has been p oposed
ha wi hin his ne wo k lichen species o ganize in communi ies known as pho obion -media ed guilds29. Guilds
a e g oups o lichen species ha a e ecologically connec ed by sha ing one o mo e pho obion s29,30 (Fig.1d).
Fungal species in a guild can bene i each o he by p opaga ing a common se o pa ne s, d i ing he es ablish-
men o connec ed lichen species in o new o ma ginal habi a s31, while compe ing o space and esou ces32.
Much o ou cu en unde s anding on pho obion -mycobion pa ne ships comes om s udies in ol ing a
ew lichen species o geog aphical a eas17,19,23,24,33–38. Howe e , mycobion -pho obion pa ne ships do no happen
in isola ion, hey a e pa o a la ge web o in e ac ions suppo ing he assembly and main enance o communi-
ies, and unde s anding hem equi es a sys emic app oach. Many species in e ac ion ne wo ks a e classi ied
as nes ed o modula . A modula ne wo k is made up o spa sely linked clus e s o dense subg aphs39–41. These
communi ies o modules may a ise due o e olu iona y and en i onmen al cons ain s42, o hey may ep esen
OPEN
1E olu ion o Ne wo ks Lab, Ins i u e o E olu iona y Biology (UPF-CSIC), Passeig Ma í im de la Ba celone a
37, Ba celona 08003, Spain. 2Eu opean Cen e o Li ing Technology (ECLT), Ca’ Bo acin, Do sodu o 3911,
Venezia 30123, I aly. *email: [email p o ec ed]-csic.es; se gi.[email p o ec ed]-csic.es
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common unc ional quali ies, such as g oupings o closely ela ed p o eins implica ed in cell communica ion. The
o he common s uc u al ea u e o ecological ne wo ks is nes edness43,44. Nes edness ela es o he hie a chical
o ganisa ion in he ne wo k whe e nodes display a endency o in e ac p e e en ially wi h subse s o pa ne s o
be e -connec ed nodes45. This concep is pa icula ly impo an in ecological ne wo k s udies ocusing on he
spa ial dis ibu ion o species, hei in e ac ions, and deg ee o indi idual specialisa ion.
Modula i y and nes edness a e concep ually dis inc s uc u es ha a e nega i ely co ela ed; o example,
a ne wo k wi h a high deg ee o modula i y is ypically associa ed wi h a low deg ee o nes edness. When
compa ed o null models, howe e , ce ain empi ical ne wo ks may exhibi bo h pa e ns43, sugges ing he pos-
sibili y o coexis ing nes edness and modula i y46. Recen s udies in ol ing small se s o lichen species and hei
in e ac ions ha e e ealed di e ging s uc u al pa e ns: om dis inc clus e s o s ongly in e ac ing species in
Pel ige a lichens38 o an embedded specialis -gene alis s uc u e in Neph oma33. Di e en explana o y causes
ha e been p oposed o he obse ed nes ed and modula pa e ns, including e olu iona y cons ain s38 as well
as he a ia ions in he na u e o he unde lying ecological in e ac ions33 (i.e. mu ualis ic s. non-mu ualis ic
ela ions). The gene al o ganiza ion o pho obion -mycobion associa ions and i s connec ion o guilds emains
la gely unknown.
He e, we add ess hese open ques ions by econs uc ing he pho obion -mycobion ne wo k, agg ega ing dec-
ades o esea ch using Sande s and Masumo o’s me a-s udy47. This ne wo k eco ds many obse a ions (
n=206
publica ions), highligh ing c i ical scales ha ope a e beyond he species le el, such as pho obion -media ed
guilds. We combine se e al ne wo k me ics in o de o unde s and he opological signa u e o guilds. Ou
indings show ha axonomy alone canno comp ehensi ely ecapi ula e guild opology. Ne wo k modula i y,
in pa icula , does no comple ely p edic species composi ion o guilds. The he e ogeneous s uc u e unco e ed
by ou analysis os e s obus ness, wi h keys one species unc ioning as b idges ac oss guilds and whose emo al
p omo es ne wo k agmen a ion and po en ially d i ing global ins abili y.
Resul s
De ining he global pho obion ‑mycobion associa ion ne wo k (PMAN). P e ious esea ch has
ocused on licheniza ion among ew species o mycobion s and pho obion s, p o iding an insu icien unde -
s anding o symbio ic in e ac ions a he la ges scales. Fo example, symbio ic ies could be ha e been shaped by
he p esence o addi ional in e ac ions in a la ge communi y. In his con ex , ne wo k echniques ha e been pa -
icula ly success ul in he s udy o mu ualis ic39 and an agonis ic46 ne wo ks om small o la ge spa io- empo al
scales48,49. He e, we ec ea e he la ges ne wo k o lichen symbion s o da e using he ull da a se o symbion
pai ings assembled in a ecen me a-s udy by Sande s and Masumo o47. This sys em belongs o he gene al
ab
c
d
cyanobac e ia
cephalodia
so edia
algae
ungi
elabo a ed
me aboli es
p o ec ion
agains
dessica ion,
p eda o s,
UV, e c.
Pho obion s
Mycobion
Guild 2Guild 1
P3 P4
M1
1P2P
M2 M5M3 M4
P4
M1
P1
P1
M3
M5
Figu e1. A ne wo k pe spec i e on lichen symbioses. (a) schema ic ep esen a ion o lichen symbiosis a
he ecological le el. Pho obion s (yellow) p o ide ene gy ixa ion hough elabo a ed me aboli es while he
mycobion (blue) comp ises he majo i y o he lichen body and p o ides p o ec ion o a ious en i onmen al
challenges. (b) Example lichen body, including dis inc o gans o seg ega e algal and cyanobac e ial pho obion s
(e.g. cephalodia). He e, lichen ep oduc ion akes place asexually h ough so edia, which include bo h he
algal (yellow) and ungal (blue) pa ne s in each p opagule. (c) Each indi idual lichen species (g ey boxes)
is composed by a mycobion (
Mj
) and one o mo e pho obion s (
Pi
). (d) The se o associa ions among
pho obion s and mycobion s de ines a bipa i e ne wo k, i s s uc u al analysis can e eal he p esence o
mesoscale s uc u es, like guilds, ela ed o he unde lying ecological ela ions and e olu iona y his o y.
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class o bipa i e ne wo ks, which ha e links be ween nodes o di e en ypes. Figu e2 displays he pho obion -
mycobion associa ion ne wo k (PMAN) ep esen ing his da ase , whe e blue and yellow nodes co espond o
mycobion s and pho obion s espec i ely.
The pho obion ‑mycobion ne wo k is modula and no nes ed. To alida e he p esence o s uc-
u al pa e ns, we compa ed he pho obion -mycobion ne wo k o a boo s ap model ha main ains he en i e
deg ee sequence o each compa men 41,50,51 (see “Ma e ials and me hods”). This gi es a nega i e con ol ha
can help us de e mine he signi icance o he s uc u es unde conside a ion52,53. Figu e3a shows he dis ibu ion
o modula i y alues o an ensemble o boo s ap andomiza ions (his og am wi h shaded egion) compa ed
o he eal da a se a e age modula i y (dashed e ical ed line). This sugges s ha he lichen ne wo k is highly
modula , mo e han he expec ed alue o he null models (
p
<10
−5
), consis en ly wi h a signi ican dec ease
in nes edness (
p
<10
−5
, see Fig.3b and Table2). Following s anda d analyses o da a comple eness54,55, we
s udied he modula i y and nes edness o subsamples using hal he da ase , i.e. eal ne wo ks and hei andomi-
za ions wi h hal he numbe o obse ed ne wo k links (see Fig.4). We disco e ed ha modula i y and nes ed-
ness a e main ained in bo h ci cums ances, implying ha he p esen ed pa e ns a e obus and independen o
sampling dep h.
Figu e2. Pho obion -mycobion associa ion ne wo k. Bipa i e ne wo k ep esen a ion o he ull da ase
analysed in ou s udy. In e ac ions in his ne wo k in ol e wo di e en ypes o nodes: blue nodes co espond
o mycobion s, and yellow nodes indica e pho obion species. The ne wo k consis s o 34 isola ed componen s,
he la ges o which has a e age deg ee
�k�=2.524
(see Table1). Ne wo k layou was au oma ically gene a ed
wi h he FMMM algo i hm (see “Ma e ials and me hods”).
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Di e ences be ween opological e sus axonomic signa u es o guilds. Guild o ganiza ion has
been adi ionally linked o pho obion iden i y (o “pho obion -media ed guild”), and mo e p ecisely, a he
genus le el29,33,54. Following his app oach, Fig.5a shows he PMAN whe e each mycobion is labelled using
he genus o hei closes pho obion (see “Ma e ials and me hods”). The labelled ne wo k comp ises se e al
con iguous clus e s con aining ela ed mycobion species, i.e., belonging o he same genus. He e, he ne wo k
appea s o display a highe densi y o connec ions wi hin each o he guilds han wi h ou side nodes, which sug-
ges s ha guilds display a common s uc u al signa u e based on modula i y.
We compa e opological modules o well-known guilds o s udy he link be ween axonomically-speci ied
guilds and modula i y (see “Ma e ials and me hods”). In he PMAN he e a e 56 axonomy-de ined guilds and
140 opological modules. We ind ha opological modules a e s a is ically smalle han he axonomy-de ined
guilds, wi h an a e age size o 7.61 species pe module e sus 26.11 species pe guild (
p<
10
−4
, see Fig.5b).
Smalle guilds ha e a s ong ma ching wi h opology-p edic ed modules, whe eas la ge guilds include se e al
smalle modules embedded wi hin hem (see he highligh ed cases o As e ochlo is and T ebouxia). This is com-
pu ed using Jacca d’s simila i y indices56 be ween each guild (de ined by axonomy) and each module (de ined
by opology). A guild o a module he e a e he se o ungal and pho obion species ha ha e a gi en label. The e
2.0
Nes edness (NODF)
0.0
0.1
0.2
P obabili y
2.
5
ba
0.68 0.70 0.72
Modula i y (QB)
0.0
0.1
0.2
P obabili y
Figu e3. Nes edness and modula i y in he pho obion -mycobion ne wo k. He e we show (a) modula i y
deno ed as
QB
and (b) nes edness calcula ed as he ull ne wo k NODF (see “Ma e ials and me hods”). The
empi ical alues a e shown as e ical ed dashed lines, while boo s ap null model dis ibu ions a e shown as a
his og am o p obabili y densi y ob ained om 100 independen andomiza ions o he ne wo k.
Table 1. Gene al p ope ies o he pho obion -mycobion ne wo k.
Gene al p ope ies De ini ion Value
P Numbe o Pho obion s 156
M Numbe o Mycobion s 926
S=P+M Numbe o Species 1082
I Numbe o In e ac ions 1311
C = I/(P
×
M) Connec ance 0.00906
k
Mean deg ee 2.4196
kP
Mean Pho obion deg ee 8.3910
Max(kPi )
Max Pho obion deg ee 75
kM
Mean Mycobion deg ee 1.4136
Max(kMi )
Max Mycobion deg ee 9
Table 2. S uc u al p ope ies o he pho obion -mycobion ne wo k. F om le o igh , mean be weenness
cen ali y o pho obion s and mycobion s (
BCP
and
BCM
espec i ely), modula i y (
QB
) and whole nes edness
(NODF). Null model me ics we e calcula ed om 100 independen ealiza ions o he ne wo k. P- alues we e
ob ained om - es s a is ics in no mally dis ibu ed a iables (
QB
, NODF) o Mann-Whi ney U ank non-
pa ame ic es in non-no mally dis ibu ed da a (
BCP
and
BCM
).
Cen ali y Modula i y Nes edness
BCP
p- alue
BCM
p- alue
QB
p- alue
NODF
p- alue
Da a 0.0188 – 0.0026 – 0.722 – 1.879 –
Boo s ap 0.01254 0.3435 0.0014 0.1848 0.691
<10−5
2.419
<10−5
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a e 21 guilds wi h a e age Jacca d simila i y less han 0.5, indica ing a signi ican disc epancy be ween opology
and axonomy, while he emaining 35 ha e e y simila communi ies (Fig.5c).
The lack o ma ching be ween axonomic and opological classi ica ion implies he exis ence o o he s uc-
u al pa e ns beyond he mesoscale de ined by guilds. Empi ical suppo o his hypo hesis is ound in he
in e nal s uc u al di e si y o wo majo guilds: Coccomyxa and T ebouxia. These guilds a e well es ablished in
he li e a u e29,32 and p o ide a eliable mic ocosm whose indi idual componen s been ex ensi ely cha ac e ized,
making hem an ideal es -bed o ou app oach. Figu e6a shows a schema ic ep esen a ion o guilds p oposed
by Rikkinen29, while Fig.6b, c, on he o he hand, shows he sub-ne wo ks o he same guilds wi h he mos cu -
en in o ma ion. These ep esen a ions in ui i ely show dis inc p ope ies, bu he ne wo k app oach allows
us o quan i y hese di e ences mo e p ecisely.
Guild in e connec i i y ma ches majo pho obion g oups. A a bigge scale, i is impo an o
assess whe he he e he in e connec i i y among guilds is s uc u ed o no . A e he e any non- i ial pa e ns
beyond he le el o guild o ganiza ion? We build a guild in e ac ion ne wo k (GIN) whe e nodes ep esen he
majo guilds (consis ing o a leas 5 species) and edges connec each pai o guilds ha sha e a leas one myco-
bion species (see “Ma e ials and me hods”). Figu e7 depic s he adjacency ma ix (a) and he ci cula layou
(b) o he GIN. Using adjacency as a su oga e o dis ance in his ne wo k, we can es ima e node simila i y
and lay ou guilds using he as clus e algo i hm57. The pho obion phylogeny can be pa ially eco e ed in he
GIN: se e al la ge g een algae guilds clus e oge he ( om He eochlo ella o As e ochlo is) ollowed by smalle
modules con aining some g een algae, Xan hophy es and cyanobac e ia loca ed in i s own domain. In Figu e7b,
he ne wo k ep esen a ion allows us o show addi ional in o ma ion: edge wid h is p opo ional o he numbe
o sha ed species amongs guilds, while node size is p opo ional o he numbe o species in each guild. Guilds
a e colo ed acco ding o pho obion ype, g een o g een algae, blue o cyanobac e ia and yellow o yellow-
g een algae. In ag eemen wi h ou p io clus e ing analysis, e e y cyanobac e ial guild g oups oge he excep
o Nos oc. This implies ha he la ge-scale o ganiza ion o guilds is no andom, bu a he he esul o biologi-
cal ai s o e olu iona y cons ain s ha b ing physiologically simila species close oge he in he ne wo k.
Guild and species con ibu ion o ne wo k obus ness. We in es iga e he PMAN’s abili y o wi h-
s and a ious ypes o pe u ba ion while also p o iding some opologically-based insigh s o p ese a ion
e o s h ough he iden i ica ion o keys one species and guilds. The deg ee o ne wo k agmen a ion caused
by species ex inc ion is quan i ied using global e iciency, which e lec s how cos ly i is o con ey in o ma ion
ac oss nodes. Global e iciency alls as he dis ance o a gi en node inc eases, e en ually app oaching ze o in a
ully disconnec ed sys em (see “Ma e ials and me hods”).
Figu e8a shows he e ec s o di e en s a egies o species emo al on he global e iciency o he PMAN:
andom emo al o species (blue), emo al based on deg ee (g een) and emo al based on cen ali y (yellow). The
PMAN is pa icula ly esis an o andom componen ailu e; e en a e emo ing 120 nodes a andom (abou
10% o he ne wo k), he o e all e iciency o he sys em is ba ely a ec ed. This is consis en wi h many obse a-
ions in he e ogeneous biological ne wo ks58, which con ain a backbone o nodes d i ing global connec i i y,
0.72 0.74 0.76 0.78 0.80 0.82
QB
0.00
0.01
0.02
0.03
0.04
0.05
0.06
0.07
P obabili y
Modula i y
Da a
50% Subse
Boo s ap 50% Subse
ab
1.0 1.2 1.4 1.6
Whole ne wo k NODF
0.00
0.01
0.02
0.03
0.04
0.05
P obabili y
Nes edness
Da a
50% Subse
Boo s ap 50% Subse
p<10-50 p<10-23
Figu e4. Valida ion o pa e ns in subsampled ne wo ks. Modula i y (a) and Nes edness (b) pa e ns o
subsampled ne wo ks in g een (con aining andomly hal o he epo ed in e ac ions) as well as hei edge
andomiza ion coun e pa s ( ed). Dis ibu ions shown con ain 50 independen da a poin s o he subsampling
and edge andomiza ion each. The pa e ns epo ed o he ull ne wo k o mo e modula han expec ed and
less nes ed han expec ed a e main ained, hus sugges ing ha hese a e no caused by sampling biases.

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bu mos species ha e ew connec ions and hei ex inc ion has a much mo e limi ed in luence on he sys em’s
s uc u e. In con as , he ne wo k is pa icula ly ulne able o a ge ed node emo al, whe he based on deg ee
o node cen ali y. While opological ne wo k p ope ies a e unlikely o in o m ecological damage, his so o
s udy migh help guide conse a ion s a egies ha seek o p e en agmen a ion and he buildup o ecological
damage.
Figu e8b shows a simula ed cascading ex inc ion e en , in which andom species disappea and wi h a gi en
p obabili y his e ec is p opaga ed o neighbo ing species. When he chance o p opaga ion inc eases, he
amoun o a ec ed species inc eases, eaching he o ali y o he sys em when he p obabili y is 1. This ype o
analysis is ca ied as a compa ison be ween he he eal da a se (blue) e sus an ensemble o boo s apped ne -
wo ks (o ange) ac ing as nega i e con ols. On a e age, he eal da a se is mo e obus o cascading ex inc ions
Rhizonema
Scy onema
Gloeocapsa
As e ochlo is
T ebouxia
T en epohlia
Diplosphae a
He eochlo ella
Symbiochlo is
Nos oc
Coccomyxa
Ellip ochlo is
My mecia
Chlo oidium
Topology
Taxonomy
102
Module size
p < 10-4
101
b
Guilds
Modules
As e ochlo is
T ebouxia
c
0.0
0.2
0.4
0.6
0.8
1.0
Topology
in o med
Taxonomy
in o med
Jacca d Simila i y Index
T en epohlia
Coccomyxa
a
Figu e5. Compa ing opological and axonomic signa u es o pho obion -media ed guilds. (a) Guilds
p edic ed by pho obion axonomic in o ma ion in he la ges subweb o he pho obion -mycobion associa ion
ne wo k. La ge guilds a e ep esen ed by colo , while mino guilds a e ep esen ed by g ey (clus e s wi h ewe
han i e nodes). Ne wo k layou was au oma ically gene a ed wi h he FMMM algo i hm (see “Ma e ials and
me hods”). (b) Module size dis ibu ions o wo me hods o guild alloca ion: a opological de ini ion based
on ne wo k modula i y (blue) and a axonomic de ini ion o guild membe ship based on pho obion gene a
(o ange). Taxonomic guilds a e signi ican ly la ge han he opological modules (p- alue ob ained om - es
s a is ic). (c) Topological modules a e inco po a ed in o guilds. Fo each module-guild pai , we show he
Jacca d’s simila i y index, which is he in e sec ion o e union o he wo se s o species. Some guilds (in blue)
co espond o opologically de ined modules, while o he s include se e al modules (in o ange, see ex ).
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ab
c
Nos oc
Guild
Coccomyxa
Guild
T ebouxia
Guild
Figu e6. E olu ion o pho obion -media ed guild ne wo ks. (a) Schema ic ep esen a ion o he guild
ela ionships and composi ion as p oposed by Rikkinen (adap ed om29). An upda ed iew o he T ebouxia
guild (b), as well as he Coccomyxa guild (c). Ne wo k isualiza ions we e gene a ed wi h OGDF lib a y (see
“Ma e ials and me hods”).
He eochlo ella
Symbiochlo is
My mecia
Apa ococcus
Coccomyxa
Ellip ochlo is
T en epohlia
Pseudochlo ella
Chlo oidium
T ebouxia
Diplosphae a
As e ochlo is
He e ococcus
Dilabi ilum
Pseudendoclonium
Nos oc
De
u e os ichococcus
Ch oococcidiopsis
Gloeocapsa
Rhizonema
Scy onema
T ebouxia
T en epohlia
Nos oc
Pseudochlo ella
Chlo oidium
Pseudendoclonium Apa ococcus
Gloeocapsa
Scy onem
a
Rhizonem
a
Ch oococcidiopsis
He e ococcus
Dilabi ilum
My mecia
As e ochlo is
Dip
l
osp
h
ae a
Deu e os ichococcus
Coccomyxa
Symbiochlo is
He eochlo ella
Ellip ochlo is
ab
He eochlo ella
Symbiochlo is
My mecia
Apa ococcus
Coccomyxa
Ellip ochlo is
T en epohlia
Pseudochlo ella
Chlo oidium
T ebouxia
Diplosphae a
As e ochlo is
He e ococcus
Dilabi ilum
Pseudendoclonium
Nos oc
De
u e os ichococcus
Ch oococcidiopsis
Gloeocapsa
Rhizonema
Scy onema
Figu e7. Global connec i i y be ween pho obion gene a. Using he pho obion genus as he ounda ion o
lichen guilds, we econs uc he guild ne wo k’s adjacency ma ix (a), whe e whi e squa es ep esen a leas one
mycobion sha ed by pho obion gene a. Pho obion s a e so ed using hei pai wise dis ances and as clus e
algo i hm (see “Ma e ials and me hods”). In (b) we show he co esponding ne wo k ep esen a ion, whe e
link wid hs a e p opo ional o he amoun o sha ed mycobion s be ween pho obion gene a, node sizes a e
p opo ional o he numbe o species (bo h pho obion s and mycobion s) belonging o ha guild and and a e
colo ed acco ding o hei g oup: g een o g een algae (Chlo ophy a and Cha ophy a), blue o cyanobac e ia
and yellow o yellow-g een algae (Xan hophy a). Ne wo k isualiza ions we e gene a ed wi h OGDF lib a y (see
“Ma e ials and me hods”).
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o a wide ange o alues. The modula pa e n p esen in he PMAN (which is emo ed in a boo s ap) ac s as
a i ewall, limi ing he each o ha m ul e ec s o local domains.
Some guilds and species ha e a g ea e in luence han o he s in p o iding global esilience due o hei
placemen wi hin he ne wo k as well as hei local cha ac e is ics (Fig.8c). O pa icula in e es a e he guilds
Scy onema and Gloeocapsa, which ac as b idges o he majo cyanobac e ial guild Rhizonema. In addi ion,
species wi hin he majo guilds Ellip ochlo is, Symbiochlo is and Chlo oidium ha e ela i ely high cen ali ies.
Rema kably, hese a e agmen ed guilds ( hey do no o m con inuous clus e s in Fig.5a) and hei cons i uen
species o en lie a he in e sec ion be ween o he majo guilds in he ne wo k.
Discussion
A he co e o he lichen symbiosis is he associa ion be ween wo unequal pa ne s, a ungus and a pho o oph.
The lexibili y o his associa ion is key o unde s and he esilience o lichen species o pollu ed and an opogenic
ecosys ems10 and hei capaci y o adap o new en i onmen s6. Unde s anding and documen ing single pa ne -
ships has been a majo ocus o lichen esea ch. Howe e , mycobion -pho obion pa ne ships do no happen in
isola ion, hey a e pa o a la ge web o in e ac ions ha shapes hei chances o o ma ion and success. While
much e o has been in es ed in cha ac e izing indi idual associa ions, he global ne wo k o in e ac ions ha
main ain he lichen symbiosis emains unknown. Using he mos comp ehensi e da a se o mycobion -pho o-
bion pa ne ships o da e, we add essed he mul i-scale na u e o lichen symbiosis om a ne wo k pe spec i e.
Ecological ne wo ks ha e eme ged as a powe ul ool o o malize bio ic in e ac ions, including mu ualis-
ic, an agonis ic and mo e complex ela ions59. This cha ac e is ic makes hem pa icula ly use ul o s udy he
so called pho obion -media ed guilds29: communi ies o lichens ha sha e a common se o pho obion s and
acili a e each o he ’s p opaga ion, bu also compe e o space and esou ces. Lichens depend on a se o cou-
pled ecological in e ac ions in ol ing species dispe sal, acili a ion and compe i ion, which a e in u n shaped
by guild s uc u e32. Mycobion s wi hin a guild a e able o p omo e he es ablishmen o one ano he in o new
habi a s by ex ending he ecological ange o hei pa ne s33. Fo example, spo e-p oducing lichens ha equi e
a compa ible pho obion upon ep oduc ion a e indi ec ly acili a ed by he asexual lichens wi hin hei guild
al eady es ablished in ha en i onmen 30.
P e iously, guilds ha e been de ined by pho obion axonomic iden i y, bu his cha ac e isa ion has been
cons ained by he scope o he da a se s, which ha e o en included a limi ed amoun o lichen species o geo-
g aphical loca ions17,23,32–36. An agg ega ed ne wo k pe spec i e allows us o ackle he s uc u al signa u e o
lichen guilds and p o ides a solid ounda ion o add ess how he gene al pa e ns o associa ion in he lichen
symbion ne wo k a ec i s obus ness.
Ou analysis shows ha he PMAN displays a s a is ically signi ican modula and non-nes ed o ganisa ion
when compa ed o he null model. This is consis en wi h p e ious obse a ions o Pel ige a lichens38. Fu he -
mo e, we compa e he p ocesses o guild alloca ion based on pho obion axa and opologically de ined guilds,
es ing p e-es ablished assump ions ega ding guild s uc u e and composi ion. Beyond he p oposed guild
mesoscale, we can iden i y o he ele an scales in lichen symbiosis. Small axonomy-de ined guilds usually
ma ch wi h opological modules, and a e ypically composed by a single pho obion linked o a ew mycobion s.
Ins ead, la ge guilds like As e ochlo is and T ebouxia a e a he complex communi ies wi h many in e ac ing
sub-modules. This newly epo ed scale in he PMAN sugges s ha addi ional unde lying ecological o gene a-
i e cons ain s (such as habi a ange o ai -dependen associa ions) may play an impo an ole in shaping
guild s uc u e.
ab
020406080100 120
Numbe o emo ed species
3.5
3.0
2.5
2.0
1.5
1.0
log
10
(Global E iciency)
Me hod
Random
Cen ali y
Deg ee
0.00 0.02 0.04 0.06 0.
08
Be weenness Cen ali y
0.02 0.04 0.06 0.08
BC
Scy onema
Ellip ochlo is
Pseudendoclonium
Apa ococcus
Gloeocapsa
Chlo oidium
Symbiochlo is
T en epohlia
T ebouxia
As e ochlo is
Rhizonema
My mecia
Coccomyxa
Diplosphae a
Nos oc
He eochlo ella
Deu e os ichococcus
Pseudochlo ella
He e ococcus
Da a
Real
Boo s ap
3.0 2.0 1.0 0.0
0.0
0.2
0.4
0.6
0.8
1.0
1.2
log10(p obabili y o p opaga ion)
F ac ion o su i ing species
c
Figu e8. Guild con ibu ion o ne wo k obus ness. (a) Decay o global e iciency (and obus ness) wi h
inc easing numbe o ex inc species o di e en emo al s a egies. Cen ali y-based (yellow) emo al o
species a ec s he global pa h leng hs and connec i i y mo e han deg ee-based (g een) o andom emo al
o species (blue). (b) Su i ing ac ion o species in a cascading ex inc ion e en in he eal (blue) and he
boo s apped (o ange) da a se . All simula ions include 100 independen eplica es in (a) and (b), wi h he solid
lines ep esen ing he mean alue o he dis ibu ion and he shaded a ea a ound i encompasses one s anda d
de ia ion o he sample. In (c) we show he a e age be weenness cen ali y o he species in each guild, ba s
s and o he s anda d e o in he dis ibu ion.
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Acco ding o he GIN analysis, ai -dependen associa ions may also in o m la ge-scale in e ac ions be ween
pho obion gene a. We ound ha guilds a e connec ed in a non- i ial manne consis en wi h he e olu iona y
his o y o pho obion s, sepa a ing algae and cyanobac e ia in o wo iden i iable clus e s. A possible explana ion
o unde s and hese indings migh be he unique way by which some cyanobac e ia a e lichenized. Lichens
ha bo ing cyanobac e ial and algal pa ne s seg ega e hem in he same o ganism using speci ic o gans called
cephalodia60. The use o speci ic and non-uni e sal mo phological s uc u es o cyanobac e ial symbion s poin s
a a dis inc p oximal e olu iona y o igin o hese inno a ions and highligh s he impo ance o mo phological
no el y as a d i e o he pa e ns obse ed in he GIN. Pu ing hese wo s uc u al pa e ns oge he , he PMAN
ecapi ula es e olu iona y his o y a he la ges scales, bu is inconsis en wi h axon-based guild de ini ions a
he smalles scales.
The ne wo k app oach has he po en ial o gi e insigh s in o he unde lying symbio ic associa ions. The na u e
o he lichen symbiosis, whe he mu ualis ic, an agonis ic, o somewhe e in be ween, is a subjec o deba e wi hin
he communi y61–64. Some au ho s ega d lichen symbiosis as mu ually bene icial among pa ne s36,64, while o h-
e s p opose a mo e complex ela ionship analogous o pho obion domes ica ion by mycobion s63,65. Unde he
lens o acul a i e mu ualism, symbiosis can be malleable, de ined by addi ional ac o s like species densi y66 o
he quali y o he ex e nal en i onmen 62. In ha sh condi ions, pho obion s in associa ion wi h ungi a e be e
p o ec ed om ex e nal challenges like p eda ion, UV adia ion and ex eme empe a u es67. Ye in op imal
en i onmen al condi ions he e a e oppo uni y cos s o li ing in associa ion; he pho obion migh be be e
o ee-li ing ins ead o ans e ing me aboli es o i s hos in exchange o a p o ec ion ha is no equi ed.
Theo e ical s udies ha e used di e en a gumen s o explain he p esence (o absence) o modula i y and nes -
edness in ecological ne wo ks, including ecological s abili y, spa ial cons ain s, o he s eng h o coe olu ion68–70.
E olu iona y ne wo k models ha e shown ha he opological ea u es o an ecological ne wo k de e mine i s
esilience, s abilizing s uc u al pa e ns based on he ypes o in e ac ions i shows. Fo ins ance, nes edness
is o en he key pa e n ound in mu ualis ic ne wo ks39,68, while modula s uc u es a e commonly associa ed
wi h an agonis ic ne wo ks42,68,69. Modula i y and nes edness a e nega i ely co ela ed bu hey can also co-occu
in spa sely connec ed andom ne wo ks43. This obse a ion highligh s he ema kable s uc u e o he PMAN,
which e en a low connec ance alues displays modula , non-nes ed s uc u es. An impo an aspec is ha
nes edness can also be p oduced o - ee by he e olu iona y gene a i e ules71, and i is s ill unclea i nes ed-
ness co ela es wi h s abili y acco ding o mo e ecen me ics72. Nes edness and modula i y a e no exclusi e
o pa icula in e ac ion ypes73, and hey can also coexis , occupying di e en scales in he ecological ne wo k
(e.g. modules ha a e hemsel es nes ed46,74,75) o di e en dimensions o hype g aph ecologies76. Ou esul s
sugges he p esence o non-mu ualis ic in e ac ions among lichen symbion s, whe he an agonism o acul a-
i e mu ualism42,68,69.
Ne wo k s uc u e can ha e a signi ican impac on he esilience and pe sis ence o species and
ecosys ems58,77. How pe u ba ions p opaga e in ecological ne wo ks has been he subjec o nume ous s udies,
b idging he gap be ween opology and popula ion dynamics. F agmen a ion o he ecological ne wo k can ha e
a de imen al in luence on e ec i e di e si y and dis up he lows o ma e , ene gy, and ecosys em se ices78,79.
In u n, di e si y has a bu e ing e ec in accumula ion o damage and he e ec s o pe u ba ions in ecologi-
cal sys ems80. Howe e , no all species ha e he same con ibu ion o ne wo k esilience. Remo al o cen al o
highly connec ed species can pose la ge h ea s o ne wo k in eg i y (see Figu e8c). A modula s uc u e also
impac s he sp ead o cascading ex inc ions, con ining pe u ba ions o small g oups o species69,81. The symbio ic
ne wo k p esen ed he e bene i s om bo h o hese pa e ns: hub species keep he ne wo k connec ed e en when
indi idual species a e andomly emo ed, and he highly modula s uc u e inc eases esilience by unc ioning
as a i ewall compa men alizing damage.
In conclusion, we ound e idence o he modula signa u e o guilds in he da ase o agg ega ed pho obion -
mycobion associa ions. Howe e , in 25 o he 56 guilds he e is lack o ag eemen be ween axonomically de ined
guilds and opological modules, e ealing a weal h o s uc u al pa e ns ope a ing a mul iple scales. This s ands
in s a k con as o he la ges pa e ns o in e ac ion in he ne wo k, which closely ollow majo e olu iona y
g oups. Ou analysis shows ha some guilds ha e a g ea e in luence han o he s in e ms o ne wo k obus ness.
Fu u e wo k will ha e o add ess he biological easons explaining he o igin o he epo ed s uc u al pa e ns
in symbion ne wo ks. Inco po a ion o opological pa e ns in heo e ical models will imp o e ou knowledge
abou symbio ic ne wo k obus ness, which is essen ial o an icipa e biodi e si y losses and ex inc ion cascades.
Ma e ials and me hods
Da a se . To build he bipa i e ne wo k o pho obion -mycobion associa ions, we use he da a om a ecen
me a-s udy47. This wo k compiles o e 200 publica ions in he ield o lichen symbiosis, desc ibing na u al in e -
ac ions be ween symbion s. The da a was supplemen ed wi h addi ional well-known in e ac ions (Loba ia’s
cyanobion pa ne s) ha we e missing in he o iginal da a se . The me a-s udy by Sande s and Ma sumo o
ocuses on publica ions a e 1988 Tsche mak-Woess’ in luen ial e iew and especially hose publica ions p o-
iding molecula e idence o species iden i y. This da ase is, howe e , biased owa ds geog aphical loca ions in
he wes e n wo ld (namely, Eu ope and No h Ame ica). Mos symbion pai s a e gene ically alida ed h ough
sequencing e o s and co espond o a ecen wa e o publica ions in he ield, bu he ea lies publica ions in
he da a se (e.g. “A monog aphy on algal cul u e” by Choda , 1913) do no p o ide he same deg ee o alida ion.
Thus, he da a se con ains some degene a e in o ma ion in which, symbion s a e iden i ied only up o he genus
le el ins ead o he species le el (
≈30
% o he da a). Simila pa e ns as hose epo ed he e we e ound in he
analyses pe aining he non-degene a e ne wo k, as well as hose limi ed o he la ges subweb.
F om he lis o na u ally obse ed associa ions, we can de ine a pho obion -mycobion associa ion ne wo k
(PMAN)
G=(P,M,E)
in e ms o he wo disjoin se s o pho obion P and mycobion M species and ano he