FEMS Microbiology Ecology 28 (1999) 301^313
MiniReview
Thioploca spp.: ¢lamentous sulfur bacteria with nitrate vacuoles
Bo Barker JÖrgensen a; *, Victor A. Gallardo
b
a
b
Max Planck Institute for Marine Microbiology, Celsiusstr. 1, D-28359 Bremen, Germany
Departamento de Oceanograf|èa, Universidad de Concepcioèn, Casilla 2407, Concepcioèn, Chile
Received 9 March 1998; received in revised form 9 November 1998; accepted 21 November 1998
Abstract
Thioploca spp. are multicellular, filamentous, colorless sulfur bacteria inhabiting freshwater and marine sediments. They
have elemental sulfur inclusions similar to the phylogenetically closely related Beggiatoa, but in contrast to these they live in
bundles surrounded by a common sheath. Vast communities of large Thioploca species live along the Pacific coast of South
America and in other upwelling areas of high organic matter sedimentation with bottom waters poor in oxygen and rich in
nitrate. Each cell of these thioplocas harbors a large liquid vacuole which is used as a storage for nitrate with a concentration of
up to 500 mM. The nitrate is used as an electron acceptor for sulfide oxidation and the bacteria may grow autotrophically or
mixotrophically using acetate or other organic molecules as carbon source. The filaments stretch up into the overlying
seawater, from which they take up nitrate, and then glide down 5^15 cm deep into the sediment through their sheaths to oxidize
sulfide formed by intensive sulfate reduction. New major occurrences have been found in recent years, both in lakes and in the
ocean, and have stimulated the interest in these fascinating bacteria. z 1999 Federation of European Microbiological
Societies. Published by Elsevier Science B.V. All rights reserved.
Keywords : Thioploca; Sul¢de-oxidizing bacteria; Nitrate respiration ; Phylogeny of sulfur bacteria ; Physiology
1. Introduction
The ¢lamentous, colorless sulfur bacteria, Thioploca spp., were ¢rst described by the German botanist
R. Lauterborn in 1907 [1]. He found bundles of these
conspicuous ¢laments contained in gelatinous
sheaths to be abundant in the mud at 15^20 m depth
in Lake Constance at the German-Swiss border and
he named them according to their appearance: thion
* Corresponding author. Tel.: +49 (421) 2028-602;
Fax: +49 (421) 2028-690; E-mail: [email protected]
for sulfur and ploka for braid. The bacteria had an
appearance similar to Beggiatoa arachnoidea: a uniseriate ¢lament with distinct crosswalls, consisting of
a row of cylindrical, disk-shaped cells with diameters
of 5^9 Wm and with many sulfur globules. In contrast to Beggiatoa, the ¢laments often had tapered
ends and occurred as bundles surrounded by a common sheath. When many ¢laments grew intertwined
in a sheath they had the appearance under the microscope of a braid. Lauterborn named the type
species Thioploca schmidlei, after his colleague and
friend, Prof. W. Schmidle.
Four years later, the Russian microbiologist S.M.
Wislouch (Visloukh) found similar but narrower (2^
4.5 Wm) ¢laments in sheaths, as he was studying mud
0168-6496 / 99 / $20.00 ß 1999 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.
PII: S 0 1 6 8 - 6 4 9 6 ( 9 8 ) 0 0 1 2 2 - 6
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samples from the Neva river near St. Petersburg [2].
He described these as a new species, Thioploca ingrica [3], named after the local geographical region,
Ingrien. An intensive survey of benthic communities
of sulfur bacteria, published by F. Koppe in 1922 [4],
showed that thioplocas at that time occurred in several lakes of northern Germany. From Lake Constance he described a third species, Thioploca minima, with only 1^2 Wm diameter. He also noted that
¢laments of di¡erent diameters could occur in the
same sheath and erroneously gave this supposedly
variable species a new name, Thioploca mixta [4,5].
What he had observed was, however, probably a
mixture of two species sharing the same sheath. A
much later search for thioplocas in Germany by S.
Maier and W.C. Preissner in 1976 [6] indicated that
these bacteria had disappeared again from many of
the old type localities, but new occurrences, e.g. in
Lake Erie in the USA, have been found.
New interest in the research on Thioploca arose
after the widespread occurrence of very large marine
Thioploca species on the Paci¢c continental shelf of
South America was published [7]. These communities
had been noticed already many years before by V.A.
Gallardo [8] as well as by G.T. Rowe and J. Waterbury, and the slimy material, often trapped in bottom trawls, was known among the local ¢shermen as
estopa (Spanish for uncleansed wool or £ax). Their
true bacterial origin as giant thioplocas was, however, realized only in 1975 when they were presented
to the microbiologist M. Shilo, then visiting Woods
Hole.
303
2. Marine thioplocas
The marine thioplocas discovered in the Paci¢c
were unique in having diameters ranging from 15
to 40 Wm and reaching lengths of many cm (Fig.
1). They are thus amongst the largest bacteria known
[10]. These marine Thioploca spp. occur in the shelf
sediments along the Paci¢c coast of South America
in masses (wet weight including sheath) of up to 1 kg
m32 . They have now been reported from many areas
of the oxygen minimum zone underlying the PeruChile Subsurface Countercurrent system at 40^280 m
water depth, a coastline of more than 3000 km and
an area exceeding 10 000 km2 . This is probably the
largest community of visible bacteria in the world.
According to their diameters (Fig. 2), the marine
thioplocas fall into several species of which two are
considered valid today [11]: the 12^20 Wm wide Thioploca chileae and the 30^43 Wm wide Thioploca
araucae (found near the Gulf of Arauco). A narrower form called Thioploca marina of 2.5^5 Wm diameter also occurs commonly on the Chilean shelf,
but it is not yet a recognized species. Occasionally,
very wide and previously undescribed Thioploca-like
¢laments of up to 125 Wm are found among these
communities (H.N. Schulz and B.B. JÖrgensen, unpublished observations). This diameter is comparable
to the largest Beggiatoa spp. found as thick mats or
loose masses around the hydrothermal vents of the
Guaymas Basin [12]. Although a taxonomy based on
diameters, similar to that used for Beggiatoa spp., is
certainly not satisfactory, it appears that the diame-
6
Fig. 1. Marine thioplocas from the shelf o¡ the coast of Chile near Concepcioèn sampled at 50^100 m water depth during summer when
the water column over the sea £oor was anoxic. A: Sediment core of 8 cm diameter showing the whitish sheaths of thioploca (`spaghetti
bacteria') extending vertically down from the surface to many cm depth. At the surface a layer of freshly deposited phytodetritus gives
the otherwise black sediment a brownish color. B: Thioploca in their transparent sheaths sieved and washed from a sediment sample. The
frame is 8 mm wide. C : Thioploca ¢laments extending up from the sediment surface and into the anoxic but nitrate containing £ow of
seawater. The frame is 15 mm wide. D: Thioploca araucae ¢lament showing the dense globules of elemental sulfur and the tapered ¢lament end which morphologically often distinguishes Thioploca spp. from Beggiatoa spp. Scale bar is 40 Wm. E: Light micrograph of sediment with Thioploca ¢laments of the two genera, T. chileae (18 Wm wide) and T. araucae (35 Wm wide). The thin ¢laments are sulfate reducing bacteria of the genus Desulfonema. An 6-shaped nematode near the center of the picture shows for comparison the giant size of
the thioplocas. F : Bundle of T. araucae extending out of their sheath. The appearence of a braid is seen where the ¢laments cross over,
hence the name Thioploca=sulfur braid. G: Confocal laser scanning micrograph of empty Thioploca sheath covered by ¢lamentous sulfate
reducing bacteria, Desulfonema sp. A non-speci¢c £uorescent stain shows how the bacteria are longitudinally oriented on the surface of
the sheath. The ¢eld is 225 Wm wide. H: Transmission electron micrograph of longitudinal thin section of T. chileae in a sheath. The ¢laments have thin cross-walls and nearly empty cells due to the large liquid vacuoles. In the thin peripheral layer of cytoplasm sulfur globules are stored. The ¢eld is 110 Wm wide. (Photographs by M. Huëttel (A+B+C+D), B.B. JÖrgensen (E+F), T. Neu (G) and S. Maier and
H. Voelker (H). Panels B and C are from [9], with permission.)
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ters of the Thioploca populations cluster within well
de¢ned limits and show less continuity than those of
Beggiatoa [13,14] (Fig. 2, Table 1).
2.1. Morphology
Fig. 2. The diameters of Thioploca ¢laments are used to de¢ne
the species. In a sediment sample from 40 m water depth o¡ the
Chilean coast, about 250 ¢lament diameters were measured and
their relative frequency is shown at ca. 1 Wm resolution. Among
the ¢laments living in sheaths (A), the two species, T. chileae
and T. araucae, are clearly distinguished, as are the ¢laments of
a narrower species which corresponds to T. marina. The T. marina occurred in larger numbers than indicated. Filaments were
also found in large numbers outside sheaths (B). Their size distribution partly di¡ered from that of the thioplocas, which indicated that these were now just thioploca ¢laments stretching out
of their sheaths. As free living ¢laments, they may be Beggiatoa
spp., although the 30^40 Wm size class distribution resembles that
of the thioplocas. (Data from [13], with permission.)
The cell lengths of the marine Thioploca spp. are
generally 0.5^1.5 times the cell diameter and a single
¢lament may contain more than a thousand cells,
thus reaching a total length of up to 7 cm [15].
The cells are separated by septa formed by the cytoplasmic membranes and the innermost layer of the
complex, four-layered cell wall [15,16]. Numerous
sulfur inclusions are found in the cytoplasm, most
probably formed by intrusions of the outer cytoplasmic membrane such as indicated from TEM micrographs of the narrower forms of Beggiatoa and Thioploca [11,15,17]. A unique cellular structure was
observed by Maier and coworkers [11,17] by transmission electron microscopy of the large thioplocas.
Inside each cell of the ¢laments they found a central
liquid vacuole which ¢lled more than 80% of the
total cellular volume and which was bounded by a
membrane (Fig. 1H). The active cytoplasm was thus
distributed as only a thin layer along the periphery
of each cell. Similar large vacuoles were discovered
¢ve years later in the giant beggiatoas living around
hydrothermal vents [18]. Also the narrow thioplocas
appear to have many small, liquid vacuoles enclosed
by a triplet membrane of similar appearance as the
cytoplasmic membrane [15].
2.2. Physiology
The main energy and carbon metabolism of Thio-
Table 1
Characteristics of the known types of Thioploca including both valid and non-valid species
Name
Freshwater species
T. minima
T. ingrica
T. schmidlei
Marine species
T. marina
T. chileae
T. araucae
Undescribed
Diameter (Wm)
Valid species
16S rRNA sequence
Reference
1^2
2^5
5^9
no
yes
yes
no
yes
no
[3]
[2]
[1]
2.5^5
12^20
30^43
60^125
no
yes
yes
no
no
yes
yes
no
[6]
[11]
[11]
unpublished
The species de¢nition is based on diameters and on freshwater or marine habitat.
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Fig. 3. 16S rRNA distance tree of the proteobacterial Q and L subdivisions showing Thioploca, Beggiatoa and representative H2 S and sulfur oxidizing bacteria. The species of Thioploca and Beggiatoa are seen to form a phylogenetically coherent group. Sulfate reducing bacteria of the N subdivision are included as outgroup. The scalebar corresponds to 0.05 substitutions per nucleotide position. Abbreviations :
Thb. = Thiobacillus, Ect. = Ectothiorhodospira. (From [26], with permission.)
ploca spp. remains uncertain, mainly because no pure
culture has yet been obtained of these organisms
despite repeated attempts [19]. The Thioploca spp.
appear to have a lower tolerance towards oxygen
and sul¢de than Beggiatoa spp. [19]. This is in accordance with their occurrence in sediments with low
sul¢de concentration [20], as already noted by Lauterborn in 1907 [1], and in sediments underlying oxygen-depleted water. They have long been expected to
be autotrophic or mixotrophic sul¢de oxidizers, in
analogy with the morphologically very similar Beggiatoa and in accordance with their accumulation of
elemental sulfur. The suggestion of Morita and coworkers [21] that the thioplocas o¡ the coast of Chile
are methylotrophs, living on methane produced in
the sediment and seeping up from coal seams running under the sea £oor, appears to be incorrect and
due to an overinterpretation of indirect, preliminary
data. Thus, the sediments o¡ the Chilean coast,
where thioplocas grow densely, have very low production rates and concentrations of methane [20],
which could not possibly provide su¤cient organic
carbon and energy for the large bacterial populations. More careful studies on the potential substrates of the Chilean thioplocas using 14 C-labeled
substrates combined with autoradiography showed,
accordingly, that methane or methanol were not taken up, whereas there was a strong incorporation of
acetate, amino acids, bicarbonate, glucose and glycine [22]. The incorporations appeared to be depend-
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Fig. 4. Thioploca distribution and biogeochemistry during summer in the sea £oor o¡ the coast of Chile at 90 m water depth. A: Depth
distribution of Thioploca in sheaths calculated as x biovolume, i.e. mm3 biovolume per cm3 sediment. B: Depth distribution of free nitrate in the pore water compared to the 100-fold higher pool of nitrate contained in Thioploca vacuoles (sum is `total NO3
3 '). The latter
was calculated from the Thioploca biovolume multiplied by the mean nitrate concentration in the ¢laments. Note di¡erence in the two
35
NO3
SO23
3 scales. C: Rates of sulfate reduction in the sediment measured from short-term radiotracer experiments using
4 . D: Sulfate
(a) and sul¢de (4H2 S, b) in the porewater. (Data from [13,20,33], with permission.)
ent on the presence of sul¢de in the medium, which
indicates that the thioplocas may be mixotrophic
sul¢de oxidizers similar to many Beggiatoa strains
[23]. Whether they are also able to grow autotrophically like the marine beggiatoas [24], as indicated by
their incorporation of H14 CO3
3 , still needs to be demonstrated.
2.3. Phylogeny
The ¢lamentous sul¢de-oxidizing bacteria Thioploca, Beggiatoa and Thiothrix are considered to be a
related group based on their morphology and physiology and are placed together in the family Beggiatoaceae [25]. The phylogenetic relationship was recently tested by 16S rRNA sequence analysis of
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several Thioploca and Beggiatoa strains [26]. The
three Thioploca species studied, T. ingrica, T. chileae
and T. araucae, form a monophyletic group within
the Q subdivision of the Proteobacteria, closely related to the beggiatoas, whereas Thiothrix followed
a di¡erent phylogenetic lineage within the gammasubdivision (Fig. 3). The identi¢cation of Thioploca
species based on diameters (Fig. 2) was found to
provide not only a di¡erentiation of morphotypes
but indeed a separation of genospecies. The picture
is, however, far from complete and in particular a
possible close relationship between the large, vacuolated Thioploca and Beggiatoa species deserves further attention. Preliminary 16S rRNA sequence data
indicate that these may be more closely related to
each other than the large Beggiatoas are to the small
Beggiatoas (A. Teske et al., Syst. Appl. Microbiol.,
in press). Thus, although the mucus sheaths of Thioploca bundles are very conspicuous, they may re£ect only a minor biochemical and behavioral di¡erence from Beggiatoa, which also secrete a ¢ne sheath
around individual ¢laments during their gliding
movement. Similarities in morphology and biology
between the cyanobacteria, e.g. Oscillatoria or Microcoleus, and the Beggiatoaceae had since the beginning of this century led to the suggestion that the
latter were colorless, `apochlorotic' cyanobacteria
[2]. The phylogenetic analysis has, however, clearly
shown that there is not a close relation between the
¢lamentous, sul¢de-oxidizing bacteria and the cyanobacteria, which are not a subdivision of the Proteobacteria, but a separate bacterial phylum.
2.4. Liquid vacuoles
Until recently, the function and the evolutionary
niche of the giant ¢laments and their cells ¢lled with
liquid vacuoles was unknown. It was observed by
Gundersen and coworkers [27] that the morphologically similar, very large beggiatoas living on the sea
£oor around hydrothermal vents in the Guaymas
Basin built 1^3 cm thick mats with a coarse mesh
structure. This, supposedly, had the function of enabling an advective water £ow through the mesh
without blowing away the beggiatoas, as easily happens with mats of the thinner species [28]. Consequently, the hydrothermal advection could enhance
the transport of oxygen and sul¢de many-fold over a
307
pure di¡usive transport, which has been found to
otherwise limit the growth and metabolic rate of
mat-forming beggiatoas [29,30]. Larkin and Henk
[31], on the other hand, suggested that `hollowness'
was an adaptation to the large cell size, whereby
di¡usive substrate limitation due to an otherwise unfavorable surface-to-volume ratio was alleviated.
Although these suggestions are probably both correct, they missed the main secret of these organisms,
discovered only years later: the vacuoles function as
an electron acceptor reservoir, an `anaerobic lung'.
3. Thioploca community on the Chilean shelf
In 1994, a new concerted e¡ort was made to study
the Chilean Thioploca communities and their role in
the biogeochemical cycles on the South American
shelf [32]. Astonishing results came immediately as
sediment cores containing Thioploca mats were
squeezed to obtain pore water samples for chemical
analyses (Fig. 4B). As the pressure was gradually
increased, the nitrate concentration measured in the
pore water suddenly stepped up from the ambient
seawater level of 20^25 WM to 3 mM, i.e. a 100fold higher concentration [33]. What happened was
obviously that the vacuoles of Thioploca burst and
released vast amounts of nitrate. Measurements of
nitrate concentrations directly in the vacuole £uid
were subsequently done by L.P. Nielsen, who cut
pieces of a few mm length of individual ¢laments
and extracted the nitrate. The subsequent spectrophotometric analysis revealed extreme concentrations of up to 500 mM NO3
3 [32]. This is equivalent
to the molar concentration of chloride in sea water.
Elemental sulfur, stored as sulfur globules in the cytoplasm, occurred at a mean concentration of 200^
300 Wmol cm33 in the ¢laments, i.e. in similar
amounts as nitrate [20]. A novel picture of the biology of the marine thioplocas emerged from this
study, which is summarized in the following sections.
3.1. Distribution and biomass
In a transect across the Chilean shelf at 36³ south,
just north of Concepcioèn, thioplocas occurred from
about 40 m water depth to beyond the shelf break at
200 m with living biomasses peaking at 120 g wet
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Fig. 5. Three-dimensional reconstruction of Thioploca sheaths in
sediment from the Chilean coast at 40 m water depth. The sediment block of 5U2.5U1 cm was rapidly frozen, cut vertically at
100 Wm increments with a cryomicrotome, and photographed in
polarized light. From 100 sequential photographs, which had
been scanned into a computer, the 3-D image was developed. At
the sediment surface a grayish color shows a dense mat of free
¢laments of either Beggiatoa or Thioploca outside their sheaths.
Below the mat, dark strands of Thioploca sheaths extended down
through the sediment with a mostly vertical orientation. (From
[13], with permission.)
weight m32 on the mid-shelf [13]. Although about
90% of Thioploca is liquid vacuoles, the 10% of active biomass, ca. 10 g wet weight m32 , is still comparable to the total biomass of the local benthic
fauna [34]. It is also comparable to the biomass of
Beggiatoa spp., 5^20 g wet weight m32 , found in
marine sediments of the eutrophic Limfjorden [14].
Due to the intense upwelling o¡ the Chilean coast,
the overlying sea water is strongly or totally depleted
of oxygen with concentrations of 6 5 WM O2 or
6 2% air saturation. The thioplocas here build 1^2
cm thick, loose mats in the muddy sea £oor, with
their dense gelatinous sheaths consolidating the sediment surface, and with 50^500 Wm thick strands
reaching 10 cm or more down into the sediment
(Figs. 1A and 4A [13]). A three-dimensional mapping of the inhabited sheaths showed that below
the mat they form a system of preferentially vertical,
unbranched mucus tunnels allowing the gliding ¢laments to migrate from the surface and deep down
into the sediment (Fig. 5).
The thioplocas appear to move almost continuously with gliding speeds of 1^3 Wm s31 , which
may enable the bacteria to move 10 cm in a day.
The ¢laments are rather rigid and may stretch several cm up from the sediment surface into the overlying, £owing seawater where they sway back and
forth in the boundary layer £ow (Fig. 1C). Underwater video recordings from a research submersible
on the Peruvian shelf have revealed a `white lawn' of
such ¢laments densely covering the sediment (unpublished observations by M.A. Arthur, W.E. Dean, R.
Jahnke and other participants of R/V Seward Johnson cruise SJ-1092). By penetrating the di¡usive
boundary layer, the thioplocas greatly increase the
surface area available for active nitrate uptake. Accordingly, experimental studies have shown that the
nitrate uptake rate per surface area of sediment increased 10-fold when the thioplocas stretched out
from the sediment as compared to a withdrawn state
[9].
3.2. Coexistence with Beggiatoa
Although the thioplocas typically live in sheaths in
bundles ranging from a few up to a hundred ¢laments per sheath, many were found at the sediment
surface apparently without a sheath. At the Bay of
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Concepcioèn on the Chilean coast, there was a transition between an apparently pure Beggiatoa community inside the bay to a mixed community of
both genera at the entrance of the bay to pure Thioploca outside. In the mixed community it was not
possible to discriminate beggiatoas from thioplocas
(occurring outside their sheaths) by simple microscopy but only by analyzing statistically their diameter distributions. The tapered ends of ¢laments,
characteristic of Thioploca but absent in Beggiatoa,
was not a consistent character of the thioplocas.
Often two or even three well-de¢ned size classes of
Thioploca occurred within the same sheath, e.g. T.
araucae, T. chileae and T. marina together [13]. This
is a puzzling phenomenon since inhabitants in one,
unbranched sheath would be expected to represent a
clone. A possible, although uncon¢rmed, explanation is that the ¢laments, which at the sediment surface, regularly stretch far out of their own sheath,
adhere to other ¢laments and passively glide together
with these down into di¡erent sheaths when those
¢laments retract. Too little is known about the physiology of individual Thioploca species to speculate
what may be the functional relationships between
di¡erent size classes when inhabiting the same
sheath.
3.3. Biogeochemistry
Due to the extremely high phytoplankton productivity in the upwelling areas o¡ the Paci¢c coast of
South America, the organic sedimentation is correspondingly high. Carbon oxidation rates of up to 50
mmol C m32 day31 were measured in the uppermost
0^10 cm of sediment [20,33], principally as a result of
sulfate reduction under the anoxic conditions (Fig.
4C). The measured sulfate reduction rates, with peak
31
and areal rates
activities of up to 5 mM SO23
4 day
32
31
of 25 mmol m day in the upper 0^10 cm, are
among the highest found in any marine sediment.
Yet, sulfate showed little depletion in the top 10^15
cm of sediment and the produced H2 S barely accumulated to measurable concentrations (91 WM) in
the porewater (Fig. 4D). This shows that the sul¢de
reoxidation was extremely rapid and e¤cient, even
during summer in the complete absence of oxygen in
the overlying water. Consequently, nitrate in the vacuoles of thioplocas appears to be the main oxidant
309
for sul¢de and is carried down into the sediment
from the overlying water inside the vacuoles of vertically migrating thioplocas.
These results are in contrast to the ¢ndings by
Henrichs and Farrington [35] from sediments underlying the oxygen minimum zone o¡ the coast of
Peru. Here, the thioplocas were surrounded by porewater with a rather high sul¢de concentration and
their H2 S oxidation capacity was apparently insu¤ciently to keep pace with the similarly high sulfate
reduction rates [36]. However, also o¡ Peru the thioploca communities occurred under the oxygen minimum zone with 6 5 WM O2 and 35^45 WM NO3
3 in
the overlying water column [35]. The absence of oxygen and availability of nitrate thus seem to be more
important for the growth of Thioploca spp. than the
surrounding sul¢de concentration.
4. Ecology of Thioploca
4.1. Chemotaxis
Simple chemotactic responses may lead to a complex migration pattern of the whole Thioploca community, as demonstrated in intact sediment cores
maintained under in situ conditions in an anoxic
£ume [9]. The organisms show a very distinct, positive reaction to nitrate and their emergence above
the sediment surface could repeatedly be triggered by
experimental addition of nitrate to the over£owing
sea water. The thioplocas, however, react negatively
to oxygen in the sea water, but the positive response
to nitrate overrides the phobic response to oxygen at
low O2 concentrations. The organisms thus appear
to be strictly microaerophilic like the beggiatoas or
may even preferentially grow anaerobically. The thioplocas showed a positive response to low sul¢de
concentrations, 6 100 WM, but a negative response
to higher sul¢de concentrations. Generally, the thioplocas appear to be more sensitive towards oxygen
than the beggiatoas and tend to lyse some hours
after exposure to air-saturated levels of oxygen. It
is not yet clear how the combined chemotactic responses of Thioploca result in the formation of
near-vertical sheaths penetrating deep down into
the sediment and in their gliding movement from
these deep tunnels up into the £owing seawater.
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Do the ¢laments continue moving in one direction
until adverse conditions are sensed, or do they have
spontaneous reversals similar to Beggiatoa [37]?
When they stretch several cm up into the over£owing
seawater, how do the long ¢laments sense when to
stop, so that they do not lose contact with their
sheath or are £ushed away by the current? In fact,
free Thioploca ¢laments have repeatedly been observed during plankton tows in the water column
over the mats.
4.2. Ecological niche
By their oriented vertical migration and their ability to store large quantities of nitrate and sulfur in
the cells, the large thioplocas occupy an ecological
niche which is unique among prokaryotic communities. By transporting nitrate intracellularly deep
down into the anoxic sea£oor, Thioploca appears
to e¡ectively eliminate the competition from other
sul¢de oxidizing bacteria, which are unable to store
an electron acceptor for extended periods but need
concurrent access to both electron acceptor and donor in their immediate microenvironment. A similar
storage of oxygen in the vacuoles would not be possible since the lipid membranes enclosing cells and
vacuoles are permeable to gases. The thioplocas thus
commute up and down, recharging their `lungs' with
nitrate at the surface and oxidizing sul¢de at depth,
thereby storing elemental sulfur globules as an energy reserve. By their special mode of life, the thioplocas also overcome the physical limitation of substrate availability, which for the beggiatoas and
other H2 S oxidizers is determined by the gradients
of counter-di¡using sul¢de and oxygen or nitrate
[29,30]. Within the 1^2 cm thick mats of Thioploca,
the combined structures of sheaths, polychaete tubes
and pelletized sediment form a porous, spongy texture which is highly permeable to current-driven advective porewater £ow [9,38]. Thus, nitrate decreased
in the pore water from 25 WM at the sediment surface and to zero at 3^4 cm depth, so the dense bacterial community within the mat had a good access
to nitrate-rich sea water.
4.3. Nitrogen metabolism
It remains to be shown whether the nitrate, which
apparently serves as a respiratory electron acceptor,
is used for denitri¢cation or whether the thioplocas
have a dissimilatory nitrate reduction to ammonia.
Preliminary evidence points towards the latter (N.P.
Revsbech and L.P. Nielsen, unpublished results).
However, McHatton et al. [39] recently found that
also the large Beggiatoa spp. living around hydrothermal vents contained nitrate-rich vacuoles. Assays
for enzyme activity showed high activity of both
RUBISCO, the CO2 -¢xing enzyme of the Calvin
cycle, and of nitrate reductase, which led to the preliminary conclusion that these bacteria are autotrophic sul¢de oxidizers using nitrate for respiration
in a membrane-bound electron transport system. The
presence of a dissimilatory nitrate metabolism in
Beggiatoa is otherwise not clear. One strain was
found to reduce nitrate to ammonium, although it
did not grow in pure culture with nitrate as the electron acceptor [40]. Sweerts et al. [41] found that nitrate was consumed e¡ectively in a freshwater mat of
Beggiatoa, and in puri¢ed material from the mat
denitri¢cation could be demonstrated using 15 N as
a tracer. It was not proven, however, whether the
15
N2 was due to Beggiatoa
conversion of 15 NO3
3 to
or to contaminating bacteria. Evidence for nitrate
respiration has recently been found in chemoautotrophic sul¢de oxidizing bacteria living as symbionts
in marine invertebrates [42].
4.4. Sulfur metabolism
It is not clear whether and how Thioploca, packed
in their sheaths and penetrating deep into the sediment, can oxidize the ambient H2 S so e¤ciently that
the pore water sul¢de concentration is kept below
1 WM (Fig. 4D). At such a low concentration, the
di¡usion gradients and thus the di¡usive £ux of H2 S
to the sheaths are correspondingly small. Data on
the geochemical redox processes in the Chilean shelf
sediments show that oxidized iron may also play an
important role as an e¡ective scavenger of H2 S [33].
This still leaves the question of how the reduced iron
is being reoxidized at a su¤cient rate to continuously
bind the sul¢de. It is an interesting observation that
the inhabited sheaths of Thioploca are densely covered by ¢lamentous sulfate reducing bacteria of the
genus Desulfonema (Fig. 1G). These are seen microscopically as thin ¢laments, longitudinally oriented on
FEMSEC 1006 25-3-99 Cyaan Magenta Geel Zwart
B.B. JÖrgensen, V.A. Gallardo / FEMS Microbiology Ecology 28 (1999) 301^313
the outside of the sheaths, and they have been identi¢ed as Desulfonema by speci¢c probes for £uorescent in situ hybridization (M. Fukui and F. Widdel,
personal communication). The close proximity of
these sulfate reducing Desulfonema and the sul¢de
oxidizing Thioploca may help to explain how such
a rapid recycling of H2 S can take place.
5. Global occurrence
By their great extension and biomass, their highly
e¤cient nitrate scavenging, and their coupling of the
nitrogen, sulfur and carbon cycles the thioplocas
may play an important role in the biogeochemistry
of marine upwelling regions. Whether they contribute signi¢cantly to the intensive denitri¢cation in
these regions still remains to be proven. The biomass
of the Thioploca population is positively related to
high sedimentation rates of organic matter and to
oxygen depletion. Thus, the bacteria depend indirectly on the local primary productivity and the intensity of upwelling. O¡ the coast of Chile and Peru,
the Thioploca biomass was low during winter and
high during summer and early fall when organic deposition was highest and the bottom water oxygen
concentrations remained below a few percent of air
saturation [34,43,44]. In accordance with this, a general decrease in Thioploca biomass was noted during
`El Ninìo' years, when the regional wind and current
systems impeded upwelling and oxygen depletion.
Marine Thioploca communities have recently also
been found in other areas of intense upwelling and
of oxygen-poor bottom water. In the northwest Arabian Sea, the monsoon-driven upwelling combined
with the in£ow of poorly oxygenated waters from
the Red Sea creates an oxygen minimum zone at
100^1000 m water depth. Sheaths with 30^40 Wm
wide Thioploca were found abundantly in the sediment at about 400 m depth [45]. Along the southwestern coast of the African continent, Thioploca has
been found in sediments o¡ the coast of Namibia
where the highly productive upwelling ecosystem of
the Benguela Current causes oxygen depletion over
the shelf ([46] and H.N. Schulz, unpublished observations). Thioplocas have also been found to inhabit
the sea £oor around hydrothermal vents in the eastern Mediterranean Sea [47]. Further habitats contin-
311
ue to be recorded and large populations have recently also been found in lakes, e.g. the Russian
Lake Baikal [48] and the Japanese Lake Biwa [49].
As renewed interest is focused on these extraordinary
bacteria, their occurrence appears to be much more
widespread than previously thought and they probably play a signi¢cant role in the biogeochemistry of
many anoxic sediments.
Acknowledgments
We thank the following colleagues for their kind
permission to present their unpublished photographs
and results: Michael A. Arthur, Manabu Fukui,
Markus Huettel, Sigfried Maier, Thomas Neu, Lars
Peter Nielsen, Niels Peter Revsbech, Heide Schulz,
Andreas Teske and Horst Voelker. Our research on
the marine thioplocas has been funded by the Chilean FONDECYT and FONDAP Programs and by
the German Max Planck Society.
References
[1] Lauterborn, R. (1907) A new genus of sulfur bacteria (Thioploca schmidlei nov. gen. nov. spec.) (in German). Ber. Dtsch.
Bot. Ges. 25, 238^242.
[2] Wislouch, S.M. (1912) Thioploca ingrica nov. sp. (in German).
Ber. Dtsch. Bot. Ges. 30, 470^473.
[3] Maier, S. (1984) Description of Thioploca ingrica sp. nov.,
nom. rev. Int. J. Syst. Bacteriol. 34, 344^345.
[4] Koppe, F. (1922) The mud £ora of the east-holsteinian lakes
and of Lake Constance (in German). Arch. Hydrobiol. 14,
619^672.
[5] Kolkwitz, R. (1955) On the sulfur bacteria Thioploca ingrica
Wislouch (in German). Ber. Dtsch. Bot. Ges. 68, 374^380.
[6] Maier, S. and Preissner, W.C. (1979) Occurrence of Thioploca
in Lake Constance and Lower Saxony, Germany. Microb.
Ecol. 5, 117^119.
[7] Gallardo, V.A. (1977) Large benthic microbial communities in
sulphide biota under Peru-Chile Subsurface Countercurrent.
Nature 268, 331^332.
[8] Gallardo, V.A. (1963) Note on the density of the benthic
fauna and the sublittoral of northern Chile. Gayana Zool.
10, 3^15.
[9] Huettel, M., Forster, S., Kloëser, S. and Fossing, H. (1996)
Vertical migration in the sediment-dwelling sulfur bacteria
Thioploca spp. in overcoming di¡usion limitations. Appl. Environ. Microbiol. 62, 1863^1872.
[10] Angert, E.R., Clements, K.D. and Pace, N.R. (1993) The
largest bacterium. Nature 362, 239^241.
FEMSEC 1006 25-3-99 Cyaan Magenta Geel Zwart
312
B.B. JÖrgensen, V.A. Gallardo / FEMS Microbiology Ecology 28 (1999) 301^313
[11] Maier, S. and Gallardo, V.A. (1984) Thioploca araucae sp.
nov. and Thioploca chileae sp. nov. Int. J. Syst. Bacteriol.
34, 414^418.
[12] Jannasch, H.W., Nelson, D.C. and Wirsen, C.O. (1989) Massive natural occurrence of unusually large bacteria (Beggiatoa
sp.) at a hydrothermal deep-sea vent site. Nature 342, 834^
836.
[13] Schulz, H.N., JÖrgensen, B.B., Fossing, H.A. and Ramsing,
N.B. (1996) Community structure of ¢lamentous, sheathbuilding sulfur bacteria, Thioploca spp., o¡ the coast of Chile.
Appl. Environ. Microbiol. 62, 1855^1862.
[14] JÖrgensen, B.B. (1977) Distribution of colorless sulfur bacteria
(Beggiatoa spp.) in a coastal marine sediment. Mar. Biol. 41,
19^28.
[15] Maier, S. and Murray, R.G.E. (1965) The ¢ne structure of
Thioploca ingrica and a comparison with Beggiatoa. Can.
J. Microbiol. 11, 645^656.
[16] Strohl, W.R., Ge¡ers, I. and Larkin, J.M. (1981) Structure of
the sulfur inclusion envelopes from four Beggiatoas. Curr.
Microbiol. 6, 75^79.
[17] Maier, S., Voëlker, H., Beese, M. and Gallardo, V.A. (1990)
The ¢ne structure of Thioploca araucae and Thioploca chileae.
Can. J. Microbiol. 36, 438^448.
[18] Nelson, D.C., Wirsen, C.O. and Jannasch, H.W. (1989) Characterization of large, autotrophic Beggiatoa spp. abundant at
hydrothermal vents of the Guaymas Basin. Appl. Environ.
Microbiol. 55, 2909^2917.
[19] Maier, S. (1980) Growth of Thioploca ingrica in a mixed culture system. Ohio J. Sci. 80, 30^32.
[20] Ferdelman, T.G., Lee, C., Pantoja, S., Harder, J., Bebout,
B.M. and Fossing, H. (1997) Sulfate reduction and methanogenesis in a Thioploca-dominated sediment o¡ the coast of
Chile. Geochim. Cosmochim. Acta, 61, 3065^3079.
[21] Morita, R.Y., Iturriaga, R. and Gallardo, V.A. (1981) Thioploca : Methylotroph and signi¢cance in the food chain. Kieler
Meeresforsch. Sonderh. 5, 384^389.
[22] Maier, S. and Gallardo, V.A. (1984) Nutritional characteristics of two marine thioplocas determined by autoradiography.
Arch. Microbiol. 139, 218^220.
[23] Strohl, W.R. and Schmidt, T.M. (1984) Mixotrophy of the
colorless, sul¢de-oxidizing, gliding bacteria Beggiatoa and Thiothrix. In: Microbial Chemoautotrophy (Strohl, W.R. and
Tuovinen, O.H., Eds.), pp. 79^95. Ohio State University
Press, Columbus, OH.
[24] Nelson, D.C. and Jannasch, H.W. (1983) Chemoautotrophic
growth of a marine Beggiatoa in sul¢de-gradient cultures.
Arch. Microbiol. 136, 262^269.
[25] Larkin, M.J. and Strohl, W.R. (1983) Beggiatoa, Thiothrix,
and Thioploca. Annu. Rev. Microbiol. 37, 341^367.
[26] Teske, A., Ramsing, N.B., Kuëver, J. and Fossing, H. (1995)
Phylogeny of Thioploca and related ¢lamentous sul¢de-oxidizing bacteria. Syst. Appl. Microbiol. 18, 517^526.
[27] Gundersen, J.K., JÖrgensen, B.B., Larsen, E. and Jannasch,
H.W. (1992) Mats of giant sulphur bacteria on deep-sea sediments due to £uctuating hydrothermal £ow. Nature 360, 454^
455.
[28] Grant, J. and Bathmann, U.V. (1987) Swept away: Resuspen-
[29]
[30]
[31]
[32]
[33]
[34]
[35]
[36]
[37]
[38]
[39]
[40]
[41]
[42]
[43]
[44]
sion of bacterial mats regulates benthic-pelagic exchange of
sulfur. Science 236, 1472^1474.
JÖrgensen, B.B. and Revsbech, N.P. (1983) Colorless sulfur
bacteria, Beggiatoa spp. and Thiovulum spp. in O2 and H2 S
microgradients. Appl. Environ. Microbiol. 45, 1261^1270.
Nelson, D.C., JÖrgensen, B.B. and Revsbech, N.P. (1986)
Growth pattern and yield of a chemoautotrophic Beggiatoa
sp. in oxygen-sul¢de microgradients. Appl. Environ. Microbiol. 52, 225^233.
Larkin, J. and Henk, M.C. (1989) Is `hollowness' an adaptation of large prokaryotes to their largeness ? Microbios Lett.
42, 69^72.
Fossing, H., Gallardo, V.A., JÖrgensen, B.B., Huëttel, M.,
Nielsen, L.P., Schulz, H., Can¢eld, D.E., Forster, S., Glud,
R.N., Gundersen, J.K., Kuëver, J., Ramsing, N.B., Teske, A.,
Thamdrup, B. and Ulloa, O. (1995) Concentration and transport of nitrate by the mat-forming sulphur bacterium Thioploca. Nature 374, 713^715.
Thamdrup, B. and Can¢eld, D.E. (1996) Pathways of carbon
oxidation in continental margin sediments o¡ central Chile.
Limnol. Oceanogr. 41, 1629^1650.
Gallardo, V.A., Carrasco, F.D., Roa, R. and Canìete, J.I.
(1995) Ecological patterns in the benthic macrobiota across
the continental shelf o¡ central Chile. Ophelia 40, 167^188.
Henrichs, S.M. and Farrington, J.W. (1984) Peru upwelling
region sediments near 15³S. 1. Remineralization and accumulation of organic matter. Limnol. Oceanogr. 29, 1^19.
Fossing, H. (1990) Sulfate reduction in shelf sediments in the
upwelling region o¡ central Peru. Cont. Shelf Res. 10, 355^
367.
MÖller, M.M., Nielsen, L.P. and JÖrgensen, B.B. (1985) Oxygen responses and mat formation by Beggiatoa spp. Appl.
Environ. Microbiol. 50, 373^382.
Reimers, C.E. (1981) Organic matter in anoxic sediments o¡
central Peru: relations of porosity, microbial decomposition
and deformation properties. Mar. Geol. 46, 175^197.
McHatton, S.C., Barry, J.P., Jannasch, H.W. and Nelson,
D.C. (1996) High nitrate concentrations in vacuolate, autotrophic marine Beggiatoa spp. Appl. Environ. Microbiol. 62,
954^958.
Vargas, A. and Strohl, W.R. (1985) Utilization of nitrate by
Beggiatoa alba. Arch. Microbiol. 142, 279^284.
Sweerts, J.-P.R.A., de Beer, D., Nielsen, L.P., Verdouw, H.,
van den Heuvel, J.C., Cohen, Y. and Cappenberg, T.E. (1990)
Denitri¢cation by sulphur oxidizing Beggiatoa spp. mats on
freshwater sediment. Nature 344, 762^763.
Hentschel, U., Cary, S.C. and Felbeck, H. (1993) Nitrate respiration in chemoautotrophic symbionts of the bivalve Lucinoma aequizonata. Mar. Ecol. Prog. Ser. 94, 35^41.
Rosenberg, R., Arntz, W.E., de Flores, E.C., Flores, L.A.,
Carbajal, G., Finger, I. and Tarazona, J. (1983) Benthos biomass and oxygen de¢ciency in the upwelling system o¡ Peru.
J. Mar. Res. 41, 263^279.
Arntz, W.E., Tarazona, J., Gallardo, V.A., Flores, L.A. and
Salzwedel, H. (1991) Benthos communities in oxygen de¢cient
shelf and upper slope areas of the Peruvian and Chilean Paci¢c coast, and changes caused by El Ninìo. In: Modern and
FEMSEC 1006 25-3-99 Cyaan Magenta Geel Zwart
B.B. JÖrgensen, V.A. Gallardo / FEMS Microbiology Ecology 28 (1999) 301^313
Ancient Continental Shelf Anoxia (Tyson, R.V. and Pearson,
T.H., Eds.), pp. 131^154. Geological Society Special Publication, No. 58.
[45] Levin, L., Gage, J., Lamont, P., Cammidge, L., Martin, C.,
Patience, A. and Crooks, J. (1997) Infaunal community structure in a low-oxygen, organic rich habitat on the Oman continental slope, NW Arabian Sea. In: The Responses of Marine
Organisms to their Environments (Hawkins, L.E. and Hutchinson, S., Eds.), pp. 223^230. Proceedings of the 30th Marine
Biology Symposium, University of Southampton.
[46] Gallardo, V.A., Klingelhoe¡er, E., Arntz, W. and Graco, M.
(1998) First report of the bacterium Thioploca in the Benguela
313
ecosystem o¡ Namibia. J. Mar. Biol. Assoc. UK 78, 1007^
1010.
[47] Dando, P.R. and Hooper, L.E. (1996) Hydrothermal Thioploca ^ more unusual bacterial mats from Milos. BRIDGE
Newslett. 11, 45^47.
[48] Namsaraev, B.B., Dulov, L.E., Dubinina, G.A., Zemskaya,
T.I., Granina, L.Z. and Karabanov, E.V. (1994) Bacterial synthesis and destruction of organic matter in microbial mats of
Lake Baikal. Microbiology 63, 193^197.
[49] Nishino, M., Fukui, M. and Nakajima, T. (1998) Dense mats
of Thioploca, gliding ¢lamentous sulfur-oxidizing bacteria in
Lake Biwa, Central Japan. Water Res. 32, 953^957.
FEMSEC 1006 25-3-99 Cyaan Magenta Geel Zwart