A niche for cyanobacteria
containing chlorophyll d
light-harvesting pigment instead of chloro-
phyll a,the form commonly found in plants,
algae and other cyanobacteria; this means
that it depends on far-red light for photo-
synthesis. Here we demonstrate photo-
synthetic activity in Acaryochloris-like
phototrophs that live underneath minute
coral-reef invertebrates (didemnid ascidi-
ans) in a shaded niche enriched in near-
infrared light. This discovery clarifies how
these cyanobacteria are able to thrive as free-
living organisms in their natural habitat.
Acaryochloris marina was first isolated
from extracts of didemnid ascidians1,2and
was presumed to be a symbiont, like the
cyanobacterium Prochloron sp., which con-
tains chlorophyll aand b,and is found inside
didemnids3. Acaryochloris marina has been
found on red algae4and a free-living Acary-
ochloris-like organism has been discovered
in a turbid saline lake5. This indicates that
cyanobacteria containing chlorophyll dmay
be fairly widespread,yet little is known about
their habitat and ecology.
In a microphotometric survey of the
didemnid ascidians Lissoclinum patella,
he cyanobacterium known as Acaryo-
chloris marina is a unique phototroph
that uses chlorophyll d as its principal
Trididemnum paracyclops,Diplosoma similis
and Diplosoma virens, we investigated the
occurrence and distribution ofcells contain-
ing chlorophyll d (for methods, see supple-
mentary information). Prochloron, and
some unicellular cyanobacteria containing
chlorophyll a and phycobiliproteins, colon-
ized internal cavities of the didemnids, but
we found no evidence ofchlorophyll din the
ascidians.However,biofilms growing on the
underside of the didemnids contained clus-
ters of pale, greenish-yellow Acaryochloris-
absorption and fluorescence features that
were characteristic of chlorophyll d (Fig.1a,
b;and see supplementary information).
We cultured these Acaryochloris-like cells
from the biofilm. Sequence analysis of the
isolate (results not shown) indicates that the
genes that encode cells’light-harvesting pro-
tein (pcbC) and 16S ribosomal RNA corre-
spond to those of A. marina: there is 100%
identity with the 302-base-pair polymerase-
chain-reaction fragment and 99% identity
with the 392-base-pair fragment,respectively
Fibre-optic microprobe measurements7
in D. virens showed intense attenuation of
visible light. Far-red light penetrated more
efficiently through the ascidian tissue, and
was enhanced relative to the incident light
owing to light-trapping effects3,7. Under all
ascidians,visible light was strongly depleted
but there was 10–20 times more far-red light,
these had spectral
providing an ideal niche for cyanobacteria
containing chlorophyll d, which absorbs
maximally at 700–720 nm (Fig.1b).
We used variable chlorophyll fluores-
cence imaging to assess photosynthesis of
the cyanobacteria containing chlorophyll d
in their natural habitat. Maximal quantum
yields ofphotosystem II (PSII) were 0.77 and
0.59 in zones comprising Prochloron and
Acaryochloris-like cells,respectively (Fig.1c).
Maximal PSII quantum yields of 0.67–0.80
have been reported for A.marina cultures8.
As expected, the quantum yield of PSII
decreased with increasing
cells were, like Prochloron, able to sustain
high photosynthetic activity at strong light
intensity (Fig.1e).A similar light adaptation
is also evident in A.marina9.
It is an apparent paradox that Acaryo-
chloris-like cells thrive in extreme shade but
show features of adaptation to strong light.
This unusual photoacclimation reflects
the fact that they live in an environment rich
in near-infrared light and that chlorophyll d
is the main light-harvesting pigment that
drives both photosystems I and II under
We conclude that Acaryochloris-like
cyanobacteria grow in biofilms beneath
enhanced over visible light and is used for
oxygenic photosynthesis. This explains the
occurrence of epiphytic A. marina on the
underside of red algae4. Cyanobacteria that
contain chlorophyll d may thrive in other
habitats with little visible light, but further
microenvironmental controls may be
important in defining the niche of these
Michael Kühl*,Min Chen†,Peter J.Ralph‡,
Ulrich Schreiber§,Anthony W.D.Larkum†
*Marine Biological Laboratory, Institute of Biology,
University of Copenhagen, 3000 Helsingør, Denmark
†School of Biological Sciences, A08, University of
Sydney, New South Wales 2006, Australia
‡Institute for Water and Environmental Resource
Management, University of Technology Sydney,
Gore Hill, New South Wales 2065, Australia
§Julius-von-Sachs-Institut für Biowissenschaften,
Universität Würzburg, 97082 Würzburg, Germany
where far-red is
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2. Miyashita, H., Ikemoto, H., Kurano, N., Miyachi, S. &
Chihara, M.J. Phycol. 39, 1247–1253 (2003).
3. Kühl,M.& Larkum,A.W.D.in Symbiosis: Mechanisms and Model
Systems(ed.Seckbach,J.) 273–290 (Kluwer,Dordrecht,2002).
4. Murakami,A., Miyashita, H., Iseki, M.,Adachi, K. &
Mimuro, M.Science 303, 1633 (2004).
5. Miller, S. R.et al. Proc. Natl Acad. Sci. USA 102,850–855 (2005).
6. Chen, M., Hiller, R. G., Howe, C. J. & Larkum,A.W. D.
Mol. Biol. Evol. 22, 21–28 (2005).
7. Kühl, M. & Fenchel, T. Microb. Ecol. 40, 94–103 (2000).
8. Schiller, H., Senger, H., Miyashita, H., Miyachi, S. & Dau, H.
Fed. Eur. Biochem. Soc. Lett. 410, 433–436 (1997).
9. Miyashita,H. et al. Plant Cell Physiol. 38, 274–281 (1997).
Photosynth. Res. 65, 269–277 (2000).
Supplementary information accompanies this communication on
Competing financial interests: declared none.
NATURE|VOL 433|24 FEBRUARY 2005|www.nature.com/nature
(% of incident light)
PSII quantum yield
Irradiance (µmol photons m–2 s–1)
Figure 1 Distribution, spectral characteristics and photosynthesis of cells containing chlorophyll d that are associated with the didemnid
ascidian Diplosoma virens. a, Vertical section through D. virens, showing the green cells of symbiotic cyanobacterium Prochloron sp.
inside cavities, and a biofilm (white) patch of Acaryochloris-like cells growing on the underside of the ascidian (scale bar, 2 mm).
b, Top, spectral absorbance (solid lines) and ultraviolet-excited fluorescence (dashed lines) of cells from the biofilm shown in a (red
curves) and cells from an A.marina culture (blue curves).Arrow,absorption maximum of chlorophyll d.Data were normalized to the maxi-
mal absorbance and fluorescence,respectively.Bottom,spectral irradiance measured below D.virens after the biofilm had been removed,
expressed as a percentage of downwelling irradiance at the tissue surface. c, d, Images as in a, but showing the maximal photosystem-II
(PSII) quantum yield of the dark-adapted section (c) and the effective PSII quantum yield at an irradiance of 585 ?mol photons m?2s?1(d).
Both variables were scaled to the same colour gradient (0–1). e, PSII quantum yield (dashed lines) and relative rates of photosynthesis
(solid lines) as a function of irradiance in Prochloron symbionts (green) and in Acaryochloris-like cells (red),taken from areas circled in a.