Received: 22 March 2021 Revised: 21 July 2021 Accepted: 16 August 2021

DOI: 10.1002/bies.202100080

R E V I E W E S S AY
Prospects & Overviews

The enigmatic Placozoa part 1: Exploring evolutionary

controversies and poor ecological knowledge
Bernd Schierwater1 Hans-Jürgen Osigus1 Tjard Bergmann1
Neil W. Blackstone2 Heike Hadrys1 Jens Hauslage3 Patrick O. Humbert4,5
Kai Kamm1 Marc Kvansakul4,5 Kathrin Wysocki1 Rob DeSalle6

1
Institute of Animal Ecology, University of
Veterinary Medicine Hannover, Foundation, Abstract
Hannover, Germany
The placozoan Trichoplax adhaerens is a tiny hairy plate and more simply organized
2
Department of Biological Sciences, Northern
than any other living metazoan. After its original description by F.E. Schulze in 1883, it
Illinois University, DeKalb, Illinois, USA
3
Gravitational Biology, Institute of Aerospace
attracted attention as a potential model for the ancestral state of metazoan organiza-
Medicine, German Aerospace Center (DLR), tion, the “Urmetazoon”. Trichoplax lacks any kind of symmetry, organs, nerve cells, mus-
Cologne, Germany
cle cells, basal lamina, and extracellular matrix. Furthermore, the placozoan genome
4
Department of Biochemistry & Genetics, La
Trobe Institute for Molecular Science, La Trobe is the smallest (not secondarily reduced) genome of all metazoan genomes. It har-
University, Melbourne, Victoria, Australia bors a remarkably rich diversity of genes and has been considered the best living
5
Research Centre for Molecular Cancer surrogate for a metazoan ancestor genome. The phylum Placozoa presently harbors
Prevention, La Trobe University, Melbourne,
Victoria 3086, Australia three formally described species, while several dozen “cryptic” species are yet await-
6
American Museum of Natural History, New ing their description. The phylogenetic position of placozoans has recently become a
York, New York, USA
contested arena for modern phylogenetic analyses and view-driven claims. Trichoplax
Correspondence offers unique prospects for understanding the minimal requirements of metazoan ani-
Bernd Schierwater, Institute of Animal Ecology, mal organization and their corresponding malfunctions.
University of Veterinary Medicine Hannover,
Foundation, Bünteweg 17d, 30559 Hannover,
KEYWORDS
Germany.
Email: [email protected] endosymbiosis, global distribution, marine model system, Placozoa, systematics, Trichoplax,
urmetazoon

INTRODUCTION an upper epithelium faces the open water, and a layer of so-called
‘’fiber cells’’ is embedded in between the epithelia.[2] After studying Tri-
The first placozoan species, Trichoplax adhaerens (Figure 1), was dis- choplax in some detail, Karl Gottlieb Grell found that this animal cannot
covered in 1883 by the German zoologist Franz Eilhard Schulze in be grouped into any of the existing phyla and he erected the new phy-
a seawater aquarium at the Zoological Institute in Graz, Austria.[1] lum ‘‘Placozoa’’ for Trichoplax.[3] For almost half a century Placozoa was
The species name is derived from the Greek ‘‘thrix’’ (‘hair’) and ‘‘plax’’ the only monotypic animal phylum, and just recently two more placo-
(‘plate’) and the Latin “adhaerens” (sticking). Trichoplax is therefore zoan species were described.[4,5] Originally, placozoans were thought
affectionately known as the ‘‘sticky hairy plate’’, a bauplan that is by far to only occur in warm subtropical and tropical ocean waters, but we
the simplest of all animals (except for some secondarily reduced para- now predict that they also occur in moderately cool waters and that
sites). The sandwich-like placozoan bauplan is made up of two epithelia the distribution range stretches as far as 55◦ N.[6]
and non-epithelial fiber cells: A lower epithelium faces the substrate, Placozoa lack any kind of permanent symmetry (but see also[7]
and[8] ) but show a clear top-bottom polarity.[1] They have no discrete

This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License, which permits use, distribution and reproduction in any
medium, provided the original work is properly cited and is not used for commercial purposes.
© 2021 The Authors. BioEssays published by Wiley Periodicals LLC

BioEssays. 2021;2100080. wileyonlinelibrary.com/journal/bies 1 of 11

https://doi.org/10.1002/bies.202100080
2 of 11 SCHIERWATER ET AL.

With this two-part review, we will introduce the reader to the

following topics in the first part: I. diversity and distribution, II.
endosymbiosis, III. life-cycle, IV. original vs. secondary simplicity, and V.
systematic position. In part 2 we will discuss VI. mitochondrial genome
evolution, VII. genomics, VIII. Trichoplax as a model organism for
biomedical research, and IX. gravitational biology and gravity sensing.

UNEXPECTED DIVERSITY AND GLOBAL

DISTRIBUTION PATTERNS

The phylum Placozoa seems to be much more diverse than initially pre-
sumed. Since 1883 T. adhaerens has been the only formally described
and accepted species within the phylum, which therefore has been the
only monotypic animal phylum for almost one and a half century.[27]
Due to practical aspects, the only morphologically described species
T. adhaerens (sensu Schulze) has been equated in retrospect to the
F I G U R E 1 Life specimen of the placozoan Trichoplax
so-called “Grell” clone, which has been identified as the mitochon-
adhaerens (haplotype H1; “Grell clone”). Photo by Hans-Jürgen Osigus
drial 16S rDNA haplotype H1.[28,29] In brief, the placozoan “haplotype-
concept” is based on a diagnostic 16S rDNA fragment, and each pla-
organs, no defined nerve or muscle cells, no extracellular matrix nor a cozoan specimen which does not possess a 100% identical nucleotide
basal membrane.[1,9] Therefore, the nine so far identified somatic cell sequence in this locus compared to all other already known placo-
types perform all functions of nutrition uptake, stimulus perception, zoan 16S haplotypes is assigned to a new consecutive number H#
locomotion, and reproduction.[1,9–12] However, the presence of addi- (see[29] ), that is, haplotype numbers are purely descriptive numbers
tional (sub-) cell types is to be expected, and the precise distribution not mirroring genetic relationships. Only recently, two more 16S hap-
of already described cell types within the animal body requires fur- lotypes have been raised to the rank of a species, that is, Hoilungia
ther specification.[12,13] Especially the recently reported localization hongkongensis (haplotype H13[4] ) and Polyplacotoma mediterranea (hap-
of gland cells in the upper epithelium[11] calls for a revision of some lotype H0[5] ). However, more than another 20 genetically well sep-
traditional concepts of placozoan morphology. Some recent studies arated placozoan haplotypes are likewise awaiting their taxonomic
also analyzed how placozoan cells communicate (and thus coordinate) classification.[30,31] Since diagnostic morphological or ecological dif-
locally or over long distances (see e.g.[14,15] and references therein). ferences are scarce in this phylum, the description of new placozoan
In the absence of a nervous system, several signaling molecules were species is difficult and requires the application of innovative species
found to play a fundamental role in animal locomotion and/or body concepts.[4,5] Although the vast majority of placozoan 16S haplotypes
shape changes. The identified ligands range from neuropeptide-like is currently not assigned to a taxonomic rank, there is nevertheless a
proteins to the gas nitric oxide and to amino acids like D-/L-aspartate, comprehensive and well-established provisional higher classification
glycine and L-glutamate.[13,16–18] Other studies analyzed the ion chan- system in Placozoa.[29,30,32] Since 2010, this classification system com-
nel and receptor repertoire found in placozoans.[14,19–26] Without prises seven well-separated 16S clades of so far undefined taxonomic
doubt, placozoans are also an important model system to study meta- ranks. Rather surprising, despite intensive sampling efforts, no new 16S
zoan signal transduction and coordinated behavior that precedes the clade has been described for more than 10 years in Placozoa, and only
evolution of synapses and a nervous system. one species (i.e., Polyplacotoma mediterranea) currently does not fit into
Trichoplax and other placozoans represent novel model organisms the provisional placozoan “clade-system” (Figure 2). A recent study on
with tremendous potential for many areas of biological and biomed- placozoan mitogenomics[31] stressed the limitations of the short diag-
ical research. The steadily growing interest in placozoans comes as nostic 16S fragment for higher systematic approaches in placozoans
no surprise as they (i) show the simplest bauplan of all metazoan ani- and suggests the usage of mitochondrial protein sequence data for
mals, (ii) possess the smallest nuclear animal genome, (iii) harbor among the illumination of inter-clade relationships. In addition, we must also
the largest mitochondrial genomes, (iv) carry representatives of all include whole nuclear genome data if we strive for a higher-level pla-
major regulatory gene families known from humans, and (v) are highly cozoan taxonomy.[4] These data will also be helpful for defining species
amenable to experimental manipulations. What can we learn from boundaries and thus estimating the total number of placozoan species
these tiny masters of morphological simplicity? As a rule of thumb, we crawling in the oceans.
here suggest “learn the fundamentals before you analyze the byzantine Ecological modeling as well as so far conducted field sampling
complexity”. As one interesting example we will use cancer research on suggests that placozoans are out there in all oceans between 55◦ N
placozoans in space at the end of this review (see part 2, topics VIII and and 44◦ S degrees latitude, that is, from Northern Scotland to New
IX). Zealand[6,30] (Figure 3). However, field sampling of placozoans in gen-
SCHIERWATER ET AL. 3 of 11

F I G U R E 2 Placozoan phylogeny of currently known clades. The phylogenetic tree is based on complete mitochondrial genome data, which
robustly resolve relationships. It must be noted that single marker genes like the 16S rRNA fail to provide robust topologies. Image taken from[31]

eral is a difficult and time-consuming matter, mostly due to the micro- allow for life even under extreme environmental conditions in the deep
scopic size of the animals. Although there are some well-established sea. Some placozoans tolerate temperatures below 10◦ C (see[30] and
sampling methods,[33,34] none of these methods are free of sam- references therein) and we have to expect a broader distribution of pla-
pling bias and they also only partially allow a comprehensive long- cozoans, that is, outside the currently reported geographical distribu-
term monitoring of placozoans in their natural habitat. These limi- tion range (Figure 3). In addition, placozoans have also been found in
tations might explain the somewhat surprising results of a recently areas with comparatively low salinity (i.e., 20‰ in mangroves[4] ), and
published field study on placozoan diversity, which resampled several we cannot exclude that some placozoan species might tolerate even
already known haplotypes, but did not find any new haplotype.[34] lower salinities.
It also seems that the genetically most divergent haplotypes show Little is known about the ecology of placozoans. Accidental obser-
lower population densities in the field and thus are more often vations revealed a prey-predator relationship between placozoans and
overlooked. some worm-like gastropods.[33,36,37] Information on other interactions
The majority of placozoan field sampling has so far been conducted with marine organisms or on the preferred diet in the field is widely
in the 0 to 5 m water depth range,[30,35] but future sampling efforts missing.[33] With respect to the feeding behavior of placozoans,
should also target all vertical areas down to deep sea habitats. The sim- several studies reported coordinated grazing of multiple individuals,
ple feeding mode (biofilm grazers) as well as the most simple body plan and directed movement towards a food source.[38–40] Beside the
without any complex morphological or physiological structures could regular uptake of nutrition by the lower epithelium,[41,42] a sporadic
4 of 11 SCHIERWATER ET AL.

F I G U R E 3 Global distribution of placozoans. Previously published placozoan records are shown as black dots. Inferred northern and southern
distribution boundaries of placozoans, respectively, are illustrated by blue lines. Data taken from[5,6,35,52,70] and references therein

uptake of nutrition by the upper epithelium has also been reported.[43] State-of-the-art and future directions
Based on observations in the laboratory, vital and healthy placozoans
can survive starvation periods of 2 to 3 days before signs of body Three recent studies addressed these questions by analyzing near com-
degradation become visible. Besides this, our knowledge on placozoan plete genome drafts of Rickettsiales bacteria obtained from two dif-
bioenergetics and metabolism is speculation and mainly deduced from ferent strains of the placozoan host Trichoplax sp. H2 and, surpris-
their gene repertoire. ingly, from one cnidarian host.[52–54] The placozoan strain in Kamm
et al.[54] was the same that was used for the recently published host
genome assembly (H2 Panama, Atlantic Ocean),[55] while the placo-
ENDOSYMBIOSIS IN PLACOZOA—A RESEARCH zoan host in Gruber-Vodicka et al.[52] was sampled in the Pacific Ocean,
FIELD WITH POTENTIAL Hawaii. The study of Klinges et al.[53] indirectly contributed to this
question by investigating the genome of a bacterium that is frequently
Endosymbionts in placozoans have first been detected in the 1970s associated with stony corals (e.g., Acropora). This bacterium turned
by the pioneers in placozoan research, Karl Grell and Gertrud out to be closely related to the endosymbiont found in T. adhaerens.
Benwitz.[44–48] In these and later works[49] endosymbiontic bacteria For clarification, Gruber-Vodicka et al.[52] proposed a new Candidatus
have been found inside the animals’ fiber cells and also in developing taxon and named their midichloriacean endosymbiont Candidatus Grel-
oocytes.[49] It has been assumed that host and symbiont form a stable lia incantans. Likewise, Klinges et al.[53] erected a Candidatus taxon and
relationship and that the bacteria are propagated to the next gener- named the Acropora endosymbiont Candidatus Aquarickettsia rhoweri.
ation both by vegetative and sexual reproduction. The first molecular The study of Gruber-Vodicka et al.[52] also found a second, less abun-
insights into the nature of the placozoan endosymbionts were derived dant and unrelated bacterium, named Ruthmannia eludens, in the host’s
from the T. adhaerens genome project.[50] Analyses following the pub- lower epithelial cells which is not subject of the further discussion.
lication of the draft genome revealed the presence of several genome All three above studies have shown that the endosymbiotic bacte-
fragments of a bacterium that phylogenetically groups within the fam- ria possess a typical Rickettsia-like genome with reduced size (around
ily Midichloriacea of the order Rickettsiales.[51] The study of Driscoll 1.3 MB) and gene content, and because of their limited metabolic
et al.[51] pointed to a potentially single endosymbiont in Trichoplax, capacities they require a host for survival. For example, the bacteria
but the limited data left several questions open: (1) What is the prin- possess a very limited capacity to synthesize amino acids. Although
ciple nature of the relationship between host and symbiont, that is, they are capable of producing their own ATP, all possess the ATP/ADP
is the bacterium parasitic, pathogenic or endosymbiotic? (2) How sta- symporter Tlc1 that exchanges host ATP with bacterial ADP, which is
ble is the relationship, is it obligate or facultative? (3) Do all placozoan a typical rickettsial trait of energy parasitism. On the other hand, the
species carry endosymbionts? (4) Are there correlations between host endosymbionts were found to be capable of producing several impor-
and endosymbiont phylogenies? tant co-factors, like riboflavin, which are potentially beneficial for the
SCHIERWATER ET AL. 5 of 11

F I G U R E 4 16S rRNA phylogeny of the proposed endosymbiont genus “Candidatus Aquarickettsia.” Endosymbiont relationships only
sometimes reflect the phylogenetic relationships of their host. For example, the placozoan haplotypes H2 Panama (H2_PAN) and H17, as well as
H13 and H15, are closely related, and so are their endosymbionts. On the other hand, some H2 strains like H2_OKHD, H2_ROS, and the H2 strain
from Hawaii[52] harbor endosymbionts that segregate with the “uncultured bacterium clone 1301APX_F11”. That the endosymbionts of H2
“Panama” and Trichoplax adhaerens (H1) are more closely related to each other than to the endosymbiont in H2 “Hawaii” has been detailed in Kamm
et al.[54] These exceptions highlight the still poorly understood complexity of host-endosymbiont relationships in marine Metazoa. Image taken
from Klinges et al.[53]

host. In some cases, the host may even complement missing bacte- highest number of 16S sequences from placozoan rickettsial endosym-
rial genes of a particular pathway. Surprisingly, only the placozoan bionts. According to this study, all of the midichloriacean type placo-
endosymbiont of the “Panama” H2 strain and A. rhoweri of Acropora zoan endosymbionts (including Grellia) fall into the Candidatus genus
were found to be capable of producing the essential amino acids lysine Aquarickettsia, together with bacteria that are associated with protists,
and threonine and potentially providing it to their hosts,[53,54] while sponges, cnidarians or whose marker sequences have been obtained
Grellia of the Hawaiian H2 strain is missing the necessary genes.[52] from sediment samples (cf.[56] ). In several cases the phylogeny of the
Ultrastructural analyses and copy number ratios of host versus sym- placozoan host matches that of the respective symbiont, that is, closely
biont genome have repeatedly shown that the number of rickettsial related placozoans harbor closely related symbionts. Some examples
bacterial cells in a placozoan is moderate and that they are restricted can be seen in Figure 4[53] : the closely related placozoan haplotypes
to the fiber cells.[44,49,51,52,54,55] This indicates that the bacteria are H2 Panama and H17 or H13 and H15 have almost identical endosym-
effectively controlled by their placozoan hosts and there is no sign bionts. On the other hand, the endosymbiont of the H2 Panama strain
that the bacteria are pathogenic to them. Experimental tests would be is more closely related to the one in H17 or H1 (T. adhaerens) than to
needed to validate how and which metabolites are exchanged between Grellia in the Hawaiian H2 strain (comparing tree topologies in[52–54] ).
host and symbiont and whether the relationship is strictly mutualis- Even more puzzling, the endosymbiont of the Panama strain is closer
tic. It seems likely that the nature of the relationship varies with a related to A. rhoweri of Acropora than to Grellia of the placozoan H2
given environmental context (e.g., food type and availability) and even strain (comparing[52–54] ). Our interpretation is that the mutualistic
closely related endosymbionts may differ in their metabolic capacities relationships are tight, but that the endosymbionts are interchange-
(see the two different H2 endosymbionts mentioned above). Probably able. A placozoan host strain may eliminate the bacteria under certain
in contrast to placozoan endosymbionts, the related midichloriacean conditions and takes up (or gets infected by) a similar bacterium. This
bacteria in scleractinian corals appear to be tolerable parasites under also fits to our rare observation that certain placozoan strains harbor
normal conditions but become pathogenic under environmental condi- no detectable midichloriacean endosymbionts, at least not for a while
tions with high nutrient availability.[53] Here an increase in the bacteria (unpubl. observations). An alternative scenario could be that related
population results in the depletion of host resources and either directly placozoans interbreed and that mitochondria and endosymbionts are
or indirectly (increased susceptibility) leads to coral White Band Dis- differentially propagated to the next generation. Based on the amount
ease. Although a comparable phenomenon is not known from Placozoa, of shared alleles of the nuclear genome it has indeed been argued that
the occasionally observed accumulation of degenerated animals and the Panama H2 strain and T. adhaerens (H1 strain Grell) must be the
sudden extinction of an entire culture dish under laboratory conditions descendants of a hybridization event, that is, are more closely related
should be investigated in the context of the dynamics of endosymbiotic than indicated by their mitochondrial 16S haplotype.[55]
bacteria.
If the relationship between host and symbiont is stable and inter-
dependent, the phylogeny of the host should mirror that of the sym- A MYSTERY: THE LIFE-CYCLE OF PLACOZOANS
biont. In order to test this, one has to compare the tree topologies of the
phylogenetic analyses of the three above studies[52–54] since they are The life-cycle of placozoans includes two different modes of vegetative
based on overlapping datasets. We here follow the taxonomic classifi- reproduction, binary fission and swarmer budding, and occasionally
cation of Klinges et al.[53] since their phylogenetic analysis includes the also bisexual reproduction[1,9,49,57] (Figure 5). Vegetative reproduction
6 of 11 SCHIERWATER ET AL.

F I G U R E 5 The three different modes of reproduction in Placozoa. Vegetative reproduction comprises fission as well as the formation of
mobile swarmer stages. In fission the mother animal divides its body in two, sometimes three (see Figure 6) daughter individuals. In swarmer
formation mode normally a single pelagic swarmer is formed and released. The complete life-cycle remains unknown, since completion of the
sexual reproduction part has never been observed yet. For details see main text

H16 have failed so far[49] indicating that a crucial factor is missing in

laboratory cultures. From genetic evidence (i.e., allele sharing patterns)
we know that sexual reproduction is rarely but regularly used at least
by Hoilungia sp. H8 in the field.[60] As of today, not a single placozoan
specimen has been collected from the field which harbored oocytes
(ref. [35] and own observations).
Our poor knowledge on the sexual life-cycle of placozoans is
unfortunate, since it might harbor clues to better understand the
sharp controversy between the substantial genetic diversity and the
remarkable phenotypic (morphological) stasis.[4] Only for the recently
described and morphological remarkably different species, Polyplaco-
toma mediterranea, the genetic distance to all other placozoans is also
mirrored by a very distinctive habitus.[5] For all other placozoan haplo-
types (species and clades) the discrepancy between morphological and
genetic diversity calls for explanations. The most simple explanation for
F I G U R E 6 Unusual three-daughter fission of a placozoan mother resolving the “morphology vs. genetics” mystery would be to assume
specimen. This is the result of consecutive fissions even if the first ??the existence of a yet unknown, morphologically different and vari-
fission has not been completed yet (i.e., the mother and the first able life cycle stage. Such a life cycle stage could derive from sexual
daughter individual are still connected, while the mother starts the reproduction, which we do not understand yet. We can also not exclude
next division). Photo by Hans-Jürgen Osigus
the possibility that functional sexual reproduction has been secondar-
ily lost in (some) extant placozoans.
by binary (sometimes also trinary (Figure 6)) fission is the standard
mode of reproduction for placozoans in the laboratory. Mean doubling
times for a population reproducing by fission can be as short as 1 to THE SIMPLEST METAZOAN ANIMAL: ORIGINAL OR
2 days under optimal conditions. Under less optimal conditions the DERIVED SIMPLICITY?
budding of planctonic swarmers is favored. These small (50–100 µm),
spheric swarmers are produced in the upper epithelium and either A global perspective
compact or hollow.[57–59] Based on observations in the laboratory,
both forms develop directly into a normal adult placozoan. In the field, Phylogeny is largely, but not entirely a guide to determine if placo-
the swarmers are released into the open water and thus are drifted zoans show secondary simplification. For instance, a collagenous extra-
by chance to a new habitat. The sexual part of the placozoan life-cycle cellular matrix is often considered a shared derived character of the
is still widely unknown (see[49] and references therein). Under labo- Metazoa and thus a feature of the common ancestor of all animals.[61]
ratory conditions, the embryos die at the 128-cell stage in Trichoplax Under this assumption placozoans, which lack an extracellular matrix
sp. H2.[49] For all other placozoan haplotypes, except for H1, H2, and (ECM), would be considered secondarily simplified (see Box 1). Alter-
H16,[49] we have never observed sexual reproduction in lab cultures natively, the ECM would be an invention of the diploblastic animals
despite various efforts to induce it (Schierwater lab, unpublished after they split off from an early placozoan. Other hypotheses of sec-
data). Also, all attempts to complete the sexual life cycle in H1, H2, or ondary simplification, like a loss of muscle and nerve cells, could also
SCHIERWATER ET AL. 7 of 11

BOX 1

The long branch leading to Placozoa combined with short internal branches as observed in phylogenetic analyses (e.g.,[70] ) has
stimulated discussions of new evolutionary hypotheses. Some of the following aspects have already been discussed in a preprint
(https://www.biorxiv.org/content/10.1101/200972v1) but have not made it into a peer-reviewed publication.[70] The scenarios discussed
in the following section are sensitive to yet unidentified major placozoan taxa (whether extant or extinct) and life-cycle stages (see main
text) and therefore should be taken with caution. It must also be highlighted that even if there had been some degree of secondary mor-
phological simplifications during the course of placozoan evolution (e.g.,[71,72] ), this would not affect the outstanding importance of nowa-
days placozoans as a comparative model system for the early evolution of Metazoa.[27]
The “ancestral vs. derived bauplan simplicity” hypotheses
Molecular systematics have jiggled the Placozoa around into nearly any possible phylogenetic position (see main text) and thus
enforced a discussion of ancestral vs. secondarily derived simplicity of the placozoan bauplan. The “derived simplicity” hypothe-
sis postulates a recent genetic bottleneck of the global placozoan diversity with the extinction of all but one placozoan group (cf.
https://www.biorxiv.org/content/10.1101/200972v1), implying that the phylum Placozoa originally harbored a variety of diverse and
deeply branching taxonomic units over the last 600 million years. Some authors furthermore speculate that extant modern placozoans
do not represent the original placozoan bauplan but have lost a basal membrane (BM) and/or extracellular matrix (ECM).[61,73,74] Other
speculations even comprise the potential loss of a Hox gene in this context.[75] However, in the absence of any confirmed fossil records
and solid phylogenetic trees for early metazoan radiation the way from such speculations to valid scientific hypotheses in sensu stricto
seems quite far.
The alternative and traditional hypothesis, that the simplicity of the placozoan bauplan is ancestral, looks very natural from many perspec-
tives, but has been questioned by some molecular analyses. The observed short branch lengths between the different placozoan groups
have challenged the view of a long period (several hundred million years) of “morphological stasis” without species and bauplan diversifi-
cation and suggested that all observed placozoan diversity is quite young.[4,70] The complex placozoan genetic toolkit[4,50,55,76] has been
added to the discussion and led to the saying that a conservation of a primitive bauplan over such long periods is “unlikely”. On the other
hand we know, however, that if there is no competition there is no grounds for radiation. It seems conceivable that for grazers like the small
low-density placozoans, a biofilm food source was never limited over longer time periods. Other potential limitations leading to compe-
tition between placozoans we also do not know of. If so, then the driving force for radiation, that is, competition for limited resources, is
simply not given, and in such a scenario the approved simplest bauplan should outcompete a more complex bauplan.

be addressed if the relationships among the five groups at the base of proteins, characterize vertebrate striated muscles. Surprisingly, MyHC
the metazoan tree of life could be resolved. For this the five principal proteins are conserved throughout animals and are even found in
groups of animals are the bilaterians (or triploblastic animals), cnidar- some unicellular organisms.[66] Sponges clearly lack muscles, but they
ians (corals and medusozoans), ctenophores (comb jellies), poriferans express representatives of both MyHC orthology groups in various cell
(sponges), and placozoans. If sponges (e.g.,[62] ) or placozoans (cf.[63] ) types. Similarly, Dayraud et al.[67] found a ctenophore-specific duplica-
are sister to the other four groups, the observed simplicity would most tion of the MyHC gene, but only one of the resulting paralogs is associ-
likely be original. Indeed, some simple sponges[64] and particularly pla- ated with muscle cells. Possibly, the common ancestor of metazoan ani-
cozoans show overall similarity to the non-metazoan holozoans such mals possessed the molecular toolkit to build muscles, but only some
as Salpingoeca rosetta. The situation would become more complicated— animals use it to build muscles (placozoans and sponges did not).
or even highly unlikely—if ctenophores were regarded sister to the The ancestors of placozoans and sponges likely were among the
other four groups.[65] While modern ctenophores are likely a relict of first metazoan animals which evolved in the late Proterozoic. In those
a once-much-larger group, they are fairly sophisticated animals, for bacteria-rich oceans,[68] multicellular organisms that used ciliae for
example, with muscles, nerves, a mouth, and gut. To derive any of locomotion and feeding conquered a broad ecological niche, which led
the other diploblast groups from ctenophores would either require to the dawn of metazoan animals. Placozoans may have persisted rela-
modification of evolutionary theory or the assumption that the early tively unchanged, while other lineages may have evolved distinct com-
ctenophores were much more primitive/simplified, that is, to a degree plexity by a combination of sequential and parallel anagenetic evolu-
that they would not be called “ctenophores” anymore (see below). In all tion using the rich genetic toolkit from the Urmetazoan (cf.[69] ). If stem
such hypotheses games we must also pay attention to the surprisingly lineages of other major metazoan groups existed at this time, they likely
strong evidence that many complex features evolved in parallel, that is, resembled placozoans and simple sponges, as dictated by the ecology
independently in many animal groups (e.g.,[66,67] ). For instance, a core of the late Proterozoic. Thus, the bauplan of placozoans and simple
set of contractile proteins, such as type II myosin heavy chain (MyHC) sponges most likely is plesiomorphic.
8 of 11 SCHIERWATER ET AL.

F I G U R E 7 The position of Placozoa within the metazoan tree of life is under dispute. It is the sole use of molecular data which have allowed to
generate all possible scenarios, every single one with high support values. The implementation of normally highly outnumbered morphological
data, however, can have substantial impact on the outcome of phylogenetic analyses if the morphological data are slightly weighted. (from[63] )

THE PHYLOGENETIC POSITION OF PLACOZOA: A phylogenomic data will only make sense when the models and methods
ONE-OF-A-KIND BATTLE FIELD of analysis take into account compositional heterogeneity.[79] How to
model this compositional heterogeneity is a subject of intense debate
As should be evident from the previous discussion, the placement and several advances have been made on the way. Some researchers
of the phylum Placozoa in the tree of life is part of one of the more prefer to recode the 20 amino acid alphabets into six letter alphabets
contentious and active areas of phylogenetic research. It is part of what based on structure and chemical characteristics of the 20 amino acids.
we have called the “sister of all other metazoa” (SOM) problem.[63] This recoding reduces the complexity of the amino acid datasets. Other
There are five major metazoan taxa that are relevant to this question— researchers prefer to impose Bayesian mixture models to correct for
Bilateria, Cnidaria, Ctenophora, Placozoa and Porifera. The latter four the heterogeneity.
of these “taxa” are described phyla and one—Bilateria—is a complex Another complicating factor in the determination of the SOM is
but monophyletic taxon with tens of phyla. There are 105 possible that most of the recent hypotheses are based entirely on molecular
ways to arrange these five ingroup taxa with one outgroup. It is clear data, with the morphological data entering as an afterthought. Neu-
that the five taxa are monophyletic themselves, so the real SOM mann et al.[63] explored the possible role of morphology in the deter-
problem boils down to which of these 105 topologies is supported mination of the SOM and showed that relatively slight weighting of the
by the data. A survey of the literature for molecular hypotheses con- morphological data can result in alternative topologies for the SOM.
cerning the identity of the sister of all other metazoans indicates that For instance, any analyses showing Ctenophora as the SOM in a com-
Porifera was the preferred SOM until about 2011 when phylogenomic bined analysis can “flip” to Porifera or Placozoa as the SOM with rela-
approaches were initiated. As more and more molecular characters tively modest weighting of morphology (Figure 7). Since we don’t really
were added to the analyses, the “Ctenophores first” hypothesis grew in know how these two categories of data need to be dealt with in phy-
preference. The current picture of the literature is rather dichotomous logenetic analysis, Neumann et al.[63] argued for caution in interpret-
as over the past 5 years (from 2015 to 2020) about 40% of publications ing what is the SOM for now. It comes as no surprise that Ctenophora
prefer Porifera first, 40% prefer Ctenophora first and the remainder first scenarios are best achieved if systematic errors in the analyses are
present the problem as ambiguous. Since from a comparative zoology introduced.[80,81]
and evolutionary point of view, any Ctenophora first scenarios must Nielsen[82] summarized the morphological implications of both
be analytical artifacts, an interesting question arises here. Does the Porifera and Ctenophora as the SOM. Only two characters—cells with
cumulation of more and more low quality characters (base pairs or collar complex and intracellular digestion—could be interpreted as
amino acid residues) favor stochastic/random outcomes? morphological characters uniting Choanoflagellata and Porifera. He
Prior to the first molecular studies that address this problem, mor- also compared the number of shared derived characters (synapo-
phological studies all supported either Porifera or Placozoa as the morphies) supporting Porifera as SOM and Ctenophora as SOM. If
SOM. As early single gene or small multiple gene studies accumu- ctenophores branched off first, Nielsen[82] concludes, there are two
lated, Porifera appeared to be the clear “winner” in the SOM sweep- options: (1) multiple morphological features that Ctenophora share
stakes. However, with the advent of phylogenomic scale datasets, also with Placozoa and Cnidaria need to be lost, or (2) significant indepen-
Ctenophora gained support as the SOM in some studies. Another trend dent or convergent evolution (homoplasies) have evolved for a large
to note is that a good proportion of the studies tended to “deresolve” number of anatomical characters.
the SOM problem and present an unresolved tree as a hypothesis Currently, then, Placozoa are enigmatic with respect to their place-
for the metazoan topology. From a comparative morphology point of ment in the animal tree of life. Full disclosure prompts us to point out
view, it is obvious that neither Cnidaria, Ctenophora nor Bilateria are that recent work[70] suggests that placozoans are sister to Cnidaria
a viable candidate for the SOM. It is clear that the determination of but we agree with King and Rokas[83] that the uncertainty of the
the SOM is highly dependent on the dataset and the methods of analy- relationships here needs to be “embraced” at this point in time. The
sis (e.g.,[62,65,77,78] ). However, a cogent argument can be made that the grand majority of the 105 hypotheses for these taxa relationships can
SCHIERWATER ET AL. 9 of 11

be whittled away and discarded, leading us to a small number of viable 7. Zuccolotto-Arellano, J., & Cuervo-González, R. (2020). Binary fission
alternatives. The future work in phylogenetics of placozoans promises in Trichoplax is orthogonal to the subsequent division plane. Mech-
anisms of Development, 162, 103608. https://doi.org10.1016/j.mod.
some novel learned lessons and will help to progress to a sound answer.
2020.103608
8. DuBuc, T. Q., Ryan, J. F., & Martindale, M. Q. (2019). "Dorsal-ventral"
genes are part of an ancient axial patterning system: Evidence from
CONCLUSIONS Trichoplax adhaerens (Placozoa). Molecular Biology and Evolution, 36(5),
966–973. https://doi.org10.1093/molbev/msz025
The enigmatic Placozoa have been attracting a lot of attention across 9. Schulze, F. E. (1891). Über Trichoplax adhaerens. In G. Reimer (Ed.),
disciplines and the momentum keeps going. This is remarkable for var- Abhandlungen der Königlichen Preuss. Akademie der Wissenschaften zu
Berlin. (pp. 1–23). Berlin: Verlag der königlichen Akademie der Wis-
ious reasons, including (i) the fact that we know very little about the
senschaften.
placozoan life-cycle, ecology, and diversity, and (ii) the odd discussions 10. Smith, C. L., Varoqueaux, F., Kittelmann, M., Azzam, R. N., Cooper, B.,
about the phylogenetic position of Placozoa in the tree of life. Winters, C. A., Eitel, M., Fasshauer, D., & Reese, T. S. (2014). Novel
cell types, neurosecretory cells, and body plan of the early-diverging
metazoan Trichoplax adhaerens. Current Biology, 24(14), 1565–1572.
ACKNOWLEDGMENTS
https://doi.org10.1016/j.cub.2014.05.046
The authors thank Kristin Fenske for her help at all different levels 11. Mayorova, T. D., Hammar, K., Winters, C. A., Reese, T. S., & Smith,
of preparing the manuscript. The authors also thank two anonymous C. L. (2019). The ventral epithelium of Trichoplax adhaerens deploys
reviewers for a wealth of critical and very helpful comments which in distinct patterns cells that secrete digestive enzymes, mucus or
diverse neuropeptides. Biology Open, 8(8). https://doi.org10.1242/bio.
clearly helped the manuscript.
045674
12. Romanova, D. Y., Varoqueaux, F., Daraspe, J., Nikitin, M. A., Eitel, M.,
CONFLICT OF INTEREST Fasshauer, D., & Moroz, L. L. (2021). Hidden cell diversity in Placozoa:
The authors declare no conflict of interest. Ultrastructural insights from Hoilungia hongkongensis. Cell and Tissue
Research. https://doi.org10.1007/s00441-021-03459-y
13. Varoqueaux, F., Williams, E. A., Grandemange, S., Truscello, L., Kamm,
AUTHOR CONTRIBUTIONS K., Schierwater, B., Jékely, G., & Fasshauer, D. (2018). High cell diversity
All authors have provided input from their personal experience work- and complex peptidergic signaling underlie placozoan behavior. Cur-
ing with Trichoplax and written sections for the manuscript. rent Biology, 28(21), 3495–3501 e3492. https://doi.org10.1016/j.cub.
2018.08.067
14. Romanova, D. Y., Smirnov, I. V., Nikitin, M. A., Kohn, A. B., Borman, A.
DATA AVAILABILITY STATEMENT I., Malyshev, A. Y., Balaban, P. M. & Moroz, L. L. (2020). Sodium action
Data sharing is not applicable to this article as no new data were cre- potentials in placozoa: Insights into behavioral integration and evo-
ated or analyzed in this study. lution of nerveless animals. Biochemical and Biophysical Research Com-
munications, 532(1), 120–126. https://doi.org10.1016/j.bbrc.2020.08.
020
ORCID
15. Moroz, L. L., Romanova, D. Y., & Kohn, A. B. (2021). Neural versus alter-
Bernd Schierwater https://orcid.org/0000-0002-3410-3660 native integrative systems: molecular insights into origins of neuro-
Neil W. Blackstone https://orcid.org/0000-0001-7195-3237 transmitters. Philosophical Transactions of the Royal Society of London.
Rob DeSalle https://orcid.org/0000-0002-6490-7109 Series B: Biological Sciences, 376(1821), 20190762. https://doi.org10.
1098/rstb.2019.0762
16. Moroz, L. L., Romanova, D. Y., Nikitin, M. A., Sohn, D., Kohn, A. B.,
REFERENCES Neveu, E., Varoqueaux, F., & Fasshauer, D. (2020). The diversification
1. Schulze, F. E. (1883). Trichoplax adhaerens, nov. gen., nov. spec. Zoolo- and lineage-specific expansion of nitric oxide signaling in Placozoa:
gischer Anzeiger, 6, 92–97. insights in the evolution of gaseous transmission. Scientific Reports,
2. Smith, C. L., Mayorova, T. D., Winters, C. A., Reese, T. S., Leys, 10(1), 13020. https://doi.org10.1038/s41598-020-69851-w
S. P., & Heyland, A. (2021). Microscopy studies of placozoans. 17. Moroz, L. L., Sohn, D., Romanova, D. Y., & Kohn, A. B. (2020).
Methods in Molecular Biology, 2219, 99–118. https://doi.org10.1007/ Microchemical identification of enantiomers in early-branching ani-
978-1-0716-0974-3_6 mals: Lineage-specific diversification in the usage of D-glutamate
3. Grell, K. G. (1971). Trichoplax adhaerens F.E. Schulze und die Entste- and D-aspartate. Biochemical and Biophysical Research Communications,
hung der Metazoan. Naturwissenschaftliche Rundschau, 24(4), 160– 527(4), 947–952. https://doi.org10.1016/j.bbrc.2020.04.135
161. 18. Romanova, D. Y., Heyland, A., Sohn, D., Kohn, A. B., Fasshauer, D., Varo-
4. Eitel, M., Francis, W. R., Varoqueaux, F., Daraspe, J., Osigus, H. J., queaux, F., & Moroz, L. L. (2020). Glycine as a signaling molecule and
Krebs, S., Vargas, S., Blum, H., Williams, G. A., Schierwater, B., & chemoattractant in Trichoplax (Placozoa): insights into the early evo-
Worheide, G. (2018). Comparative genomics and the nature of pla- lution of neurotransmitters. Neuroreport, 31(6), 490–497. https://doi.
cozoan species. Plos Biology, 16(7), e2005359. https://doi.org10.1371/ org10.1097/wnr.0000000000001436
journal.pbio.2005359 19. Senatore, A., Raiss, H., & Le, P. (2016). Physiology and evolution
5. Osigus, H. J., Rolfes, S., Herzog, R., Kamm, K., & Schierwater, B. (2019). of voltage-gated calcium channels in early diverging animal phyla:
Polyplacotoma mediterranea is a new ramified placozoan species. Cur- Cnidaria, Placozoa, Porifera and Ctenophora. Frontiers in Physiology, 7,
rent Biology, 29(5), R148–R149. https://doi.org10.1016/j.cub.2019.01. 481. https://doi.org10.3389/fphys.2016.00481
068 20. Novotný, J. P., Chughtai, A. A., Kostrouchová, M., Kostrouchová, V.,
6. Paknia, O., & Schierwater, B. (2015). Global habitat suitability and eco- Kostrouch, D., Kaššák, F., Kaňa, R., Schierwater, B., Kostrouchová,
logical niche separation in the phylum Placozoa. Plos One, 10(11). https: M., & Kostrouch, Z. (2017). Trichoplax adhaerens reveals a network
//doi.orgARTNe014016210.1371/journal.pone.0140162 of nuclear receptors sensitive to 9-cis-retinoic acid at the base of
10 of 11 SCHIERWATER ET AL.

metazoan evolution. PeerJ, 5, e3789. https://doi.org10.7717/peerj. 38. Smith, C. L., Pivovarova, N., & Reese, T. S. (2015). Coordinated feed-
3789 ing behavior in Trichoplax, an animal without synapses. Plos One, 10(9),
21. Smith, C. L., Abdallah, S., Wong, Y. Y., Le, P., Harracksingh, A. N., Artinian, e0136098. https://doi.org10.1371/journal.pone.0136098
L., Tamvacakis, A. N., Rehder, V., Reese, T. S., & Senatore, A. (2017). 39. Smith, C. L., Reese, T. S., Govezensky, T., & Barrio, R. A. (2019). Coher-
Evolutionary insights into T-type Ca(2+) channel structure, function, ent directed movement toward food modeled in Trichoplax, a ciliated
and ion selectivity from the Trichoplax adhaerens homologue. Jour- animal lacking a nervous system. Proceedings of the National Academy
nal of General Physiology, 149(4), 483–510. https://doi.org10.1085/jgp. of Sciences of the United States of America, 116(18), 8901–8908. https:
201611683 //doi.org10.1073/pnas.1815655116
22. Elkhatib, W., Smith, C. L., & Senatore, A. (2019). A Na(+) leak chan- 40. Fortunato, A., & Aktipis, A. (2019). Social feeding behavior of Tri-
nel cloned from Trichoplax adhaerens extends extracellular pH and choplax adhaerens. Frontiers in Ecology and Evolution, 7, 19. https://doi.
Ca(2+) sensing for the DEG/ENaC family close to the base of Meta- org10.3389/fevo.2019.00019
zoa. Journal of Biological Chemistry, 294(44), 16320–16336. https://doi. 41. Smith, C. L., & Mayorova, T. D. (2019). Insights into the evolu-
org10.1074/jbc.RA119.010542 tion of digestive systems from studies of Trichoplax adhaerens.
23. Gauberg, J., Abdallah, S., Elkhatib, W., Harracksingh, A. N., Piekut, T., Cell and Tissue Research, 377(3), 353–367. https://doi.org10.1007/
Stanley, E. F., & Senatore, A. (2020). Conserved biophysical features s00441-019-03057-z
of the Ca(V)2 presynaptic Ca(2+) channel homologue from the early- 42. Smith, C. L., & Reese, T. S. (2016). Adherens junctions modulate dif-
diverging animal Trichoplax adhaerens. Journal of Biological Chemistry, fusion between epithelial cells in Trichoplax adhaerens. Biological Bul-
295(52), 18553–18578. https://doi.org10.1074/jbc.RA120.015725 letin, 231(3), 216–224. https://doi.org10.1086/691069
24. Gauberg, J., Senatore, A., & Heyland, A. (2021). Functional stud- 43. Wenderoth, H. (1986). Transepithelial cytophagy by Trichoplax
ies of Trichoplax adhaerens voltage-gated calcium channel activity. adhaerens F.E.Schulze (Placozoa) feeding on yeast. Zeitschrift für
Methods in Molecular Biology, 2219, 277–288. https://doi.org10.1007/ Naturforschung C, 41(3), 343–347.
978-1-0716-0974-3_18 44. Grell, K. G., Schulze, F. E. & Benwitz, G. (1971). Die Ultrastruktur von
25. Reitzel, A. M., Macrander, J., Mane-Padros, D., Fang, B., Sladek, F. M., & Trichoplax adhaerens. Cytobiologie, 4, 216–240.
Tarrant, A. M. (2018). Conservation of DNA and ligand binding proper- 45. Grell, K. G. (1972). Eibildung und Furchung von Trichoplax adhaerens
ties of retinoid X receptor from the placozoan Trichoplax adhaerens to F.E.Schulze (Placozoa). Zeitschrift für Morphologie der Tiere, 73, 297–
human. Journal of Steroid Biochemistry and Molecular Biology, 184, 3–10. 314.
https://doi.org10.1016/j.jsbmb.2018.02.010 46. Grell, K. G., & Benwitz, G. (1974). Elektronenmikroskopische Beobach-
26. Kuznetsov, A. V., Vainer, V. I., Volkova, Y. M., & Kartashov, L. E. (2021). tungen über das Wachstum der Eizelle und die Bildung der "Befruch-
Motility disorders and disintegration into separate cells of Trichoplax tungsmembran" von Trichoplax adhaerens F.E.Schulze (Placozoa).
sp. H2 in the presence of Zn(2+) ions and L-cysteine molecules: A Zeitschrift für Morphologie der Tiere, 79, 295–310.
systems approach. Bio Systems, 206, 104444. https://doi.org10.1016/ 47. Grell, K. G., & Benwitz, G. (1974). Spezifische Verbindungsstrukturen
j.biosystems.2021.104444 der Faserzellen von Trichoplax adhaerens F. E. Schulze. Zeitschrift für
27. Schierwater, B. (2005). My favorite animal, Trichoplax adhaerens. Naturforschung C, 29(11-12), 790.
Bioessays, 27(12), 1294–1302. https://doi.org10.1002/bies.20320 48. Grell, K. G., & Benwitz, G. (1981). Ergänzende Untersuchungen
28. Ender, A., & Schierwater, B. (2003). Placozoa are not derived Cnidari- zur Ultrastruktur von Trichoplax adhaerens F.E. Schulze (Placozoa).
ans: Evidence from molecular morphology. Molecular Biology and Evolu- Zoomorphology, 98(1), 47–67.
tion, 20(1), 130–134. https://doi.org10.1093/molbev/msg018 49. Eitel, M., Guidi, L., Hadrys, H., Balsamo, M., & Schierwater, B. (2011).
29. Voigt, O., Collins, A. G., Pearse, V. B., Pearse, J. S., Ender, A., Hadrys, H., New insights into placozoan sexual reproduction and development.
& Schierwater, B. (2004). Placozoa—No longer a phylum of one. Current Plos One, 6(5), e19639. https://doi.org10.1371/journal.pone.0019639
Biology, 14(22), R944–945. https://doi.org10.1016/j.cub.2004.10.036 50. Srivastava, M., Begovic, E., Chapman, J., Putnam, N. H., Hellsten, U.,
30. Eitel, M., Osigus, H. J., DeSalle, R., & Schierwater, B. (2013). Global Kawashima, T., Kuo, A., Mitros, T., Salamov, A., Carpenter, M. L., Sig-
diversity of the Placozoa. Plos One, 8(4), e57131. https://doi.org10. norovitch, A. Y., Moreno, M. A., Kamm, K., Grimwood, J., Schmutz, J.,
1371/journal.pone.0057131 Shapiro, H., Grigoriev, I. V., Buss, L. W., Schierwater, B., . . . Rokhsar, D. S.
31. Miyazawa, H., Osigus, H. J., Rolfes, S., Kamm, K., Schierwater, B., & (2008). The Trichoplax genome and the nature of placozoans. Nature,
Nakano, H. (2021). Mitochondrial genome evolution of placozoans: 454(7207), 955–960. https://doi.org10.1038/nature07191
Gene rearrangements and repeat expansions. Genome Biology and Evo- 51. Driscoll, T., Gillespie, J. J., Nordberg, E. K., Azad, A. F., & Sobral, B. W.
lution, 13(1), evaa213. https://doi.org10.1093/gbe/evaa213 (2013). Bacterial DNA sifted from the Trichoplax adhaerens (Animalia:
32. Eitel, M., & Schierwater, B. (2010). The phylogeography of the Placozoa Placozoa) genome project reveals a putative rickettsial endosymbiont.
suggests a taxon-rich phylum in tropical and subtropical waters. Molec- Genome Biology and Evolution, 5(4), 621–645. https://doi.org10.1093/
ular Ecology, 19(11), 2315–2327. https://doi.org10.1111/j.1365-294X. gbe/evt036
2010.04617.x 52. Gruber-Vodicka, H. R., Leisch, N., Kleiner, M., Hinzke, T., Liebeke, M.,
33. Pearse, V. B., & Voigt, O. (2007). Field biology of placozoans (Tri- McFall-Ngai, M., Hadfield, M. G., & Dubilier, N. (2019). Two intracel-
choplax): Distribution, diversity, biotic interactions. Integrative and lular and cell type-specific bacterial symbionts in the placozoan Tri-
Comparative Biology, 47(5), 677–692. choplax H2. Nature Microbiology, 4(9), 1465–1474. https://doi.org10.
34. Miyazawa, H., & Nakano, H. (2018). Multiple surveys employing a new 1038/s41564-019-0475-9
sample-processing protocol reveal the genetic diversity of placozoans 53. Klinges, J. G., Rosales, S. M., McMinds, R., Shaver, E. C., Shantz, A. A.,
in Japan. Ecology and Evolution, 8(5), 2407–2417. https://doi.org10. Peters, E. C., Eitel, M., Wörheide, G., Sharp, K. H., Burkepile, D. E.,
1002/ece3.3861 Silliman, B. R., & Vega Thurber, R. L. (2019). Phylogenetic, genomic,
35. Voigt, O., & Eitel, M. (2018). Placozoa. In A. Schmidt-Rhaesa (Ed.), Mis- and biogeographic characterization of a novel and ubiquitous marine
cellaneous Invertebrates (pp. 41–54). Berlin, Boston: De Gruyter. invertebrate-associated Rickettsiales parasite, Candidatus Aquarick-
36. Riedl, R. (1959). Beitrage zur Kenntnis der Rhodope veranii, Teil I. ettsia rohweri, gen. nov., sp. nov. The ISME Journal, 13(12), 2938–2953.
Geschichte und Biologie. Zoologischer Anzeiger, 163(3), 4. https://doi.org10.1038/s41396-019-0482-0
37. Cuervo-Gonzalez, R. (2017). Rhodope placozophagus (Heter- 54. Kamm, K., Osigus, H. J., Stadler, P. F., DeSalle, R., & Schierwater,
obranchia) a new species of turbellarian-like Gastropoda that B. (2019). Genome analyses of a placozoan rickettsial endosym-
preys on placozoans. Zoologischer Anzeiger, 270, 43–48. https://doi. biont show a combination of mutualistic and parasitic traits. Scientific
org10.1016/j.jcz.2017.09.005 Reports, 9(1), 17561. https://doi.org10.1038/s41598-019-54037-w
SCHIERWATER ET AL. 11 of 11

55. Kamm, K., Osigus, H. J., Stadler, P. F., DeSalle, R., & Schierwater, and Cnidaria in genes with minimal compositional bias. Elife, 7, e36278.
B. (2018). Trichoplax genomes reveal profound admixture and sug- https://doi.org10.7554/eLife.36278
gest stable wild populations without bisexual reproduction. Scientific 71. O’Malley, M. A., Wideman, J. G., & Ruiz-Trillo, I. (2016). Losing complex-
Reports, 8(1), 11168. https://doi.org10.1038/s41598-018-29400-y ity: The role of simplification in macroevolution. Trends in Ecology & Evo-
56. Kim, M., Oh, H. S., Park, S. C., & Chun, J. (2014). Towards a taxonomic lution, 31(8), 608–621. https://doi.org10.1016/j.tree.2016.04.004
coherence between average nucleotide identity and 16S rRNA gene 72. Adamska, M. (2016). Sponges as models to study emergence of com-
sequence similarity for species demarcation of prokaryotes. Interna- plex animals. Current Opinion in Genetics & Development, 39, 21–28.
tional Journal of Systematic and Evolutionary Microbiology, 64(Pt 2), 346– https://doi.org10.1016/j.gde.2016.05.026
351. https://doi.org10.1099/ijs.0.059774-0 73. Dohrmann, M., & Worheide, G. (2013). Novel scenarios of early animal
57. Thiemann, M., & Ruthmann, A. (1988). Trichoplax adhaerens Schulze, evolution-is it time to rewrite textbooks? Integrative and Comparative
F. E. (Placozoa) - The formation of swarmers. Zeitschrift für Natur- Biology, 53(3), 503–511. https://doi.org10.1093/icb/ict008
forschung C, 43(11-12), 955–957. 74. Warren, C. R., Kassir, E., Spurlin, J., Martinez, J., Putnam, N. H., &
58. Thiemann, M., & Ruthmann, A. (1990). Spherical forms of Trichoplax Farach-Carson, M. C. (2015). Evolution of the perlecan/HSPG2 gene
adhaerens. Zoomorphology, 110(1), 37-45. and its activation in regenerating Nematostella vectensis. Plos One,
59. Thiemann, M., & Ruthmann, A. (1991). Alternative modes of asex- 10(4), e0124578. https://doi.org10.1371/journal.pone.0124578
ual reproduction in Trichoplax-adhaerens (Placozoa). Zoomorphology, 75. Ferrier, D. E. (2016). The origin of the Hox/ParaHox genes, the Ghost
110(3), 165–174. https://doi.org10.1007/Bf01632872 Locus hypothesis and the complexity of the first animal. Brief Funct
60. Signorovitch, A. Y., Dellaporta, S. L., & Buss, L. W. (2005). Molecular sig- Genomics, 15(5), 333–341. https://doi.org10.1093/bfgp/elv056
natures for sex in the Placozoa. Proceedings of the National Academy of 76. Belahbib, H., Renard, E., Santini, S., Jourda, C., Claverie, J. M.,
Sciences of the United States of America, 102(43), 15518–15522. https: Borchiellini, C., & Le Bivic, A. (2018). New genomic data and anal-
//doi.org10.1073/pnas.0504031102 yses challenge the traditional vision of animal epithelium evolution.
61. Ax, P. (1996). Multicellular Animals—A new Approach to the Phylogenetic Bmc Genomics [Electronic Resource], 19(1), 393. https://doi.org10.1186/
Order in Nature Volume, 1: Springer-Verlag Berlin Heidelberg. s12864-018-4715-9
62. Simion, P., Philippe, H., Baurain, D., Jager, M., Richter, D. J., Di Franco, 77. Dunn, C. W., Hejnol, A., Matus, D. Q., Pang, K., Browne, W. E., Smith, S.
A., Roure, B., Satoh, N., Quéinnec, É., Ereskovsky, A., Lapébie, P., Corre, A., Seaver, E., Rouse, G. W., Obst, M., Edgecombe, G. D., Sørensen, M.
E., Delsuc, F., King, N., Wörheide, G., & Manuel, M. (2017). A large and V., Haddock, S. H. D., Schmidt-Rhaesa, A., Okusu, A., Kristensen, R. M.,
consistent phylogenomic dataset supports sponges as the sister group Wheeler, W. C., Martindale, M. Q., & Giribet, G. (2008). Broad phyloge-
to all other animals. Current Biology, 27(7), 958-967. https://doi.org/10. nomic sampling improves resolution of the animal tree of life. Nature,
1016/j.cub.2017.02.031 452(7188), 745–749. https://doi.org10.1038/nature06614
63. Neumann, J. S., DeSalle, R., Narechania, A., Schierwater, B., & Tessler, 78. Whelan, N. V., Kocot, K. M., Moroz, T. P., Mukherjee, K., Williams, P.,
M. (2020). Morphological characters can strongly influence early ani- Paulay, G., Moroz, L. L., & Halanych, K. M. (2017). Ctenophore rela-
mal relationships inferred from phylogenomic datasets. Systematic tionships and their placement as the sister group to all other animals.
Biology. 70, 360–375. https://doi.org/10.1093/sysbio/syaa038 Nature Ecology & Evolution, 1(11), 1737–1746. https://doi.org10.1038/
64. Laundon, D., Larson, B. T., McDonald, K., King, N., & Burkhardt, P. s41559-017-0331-3
(2019). The architecture of cell differentiation in choanoflagellates 79. Feuda, R., Dohrmann, M., Pett, W., Philippe, H., Rota-Stabelli, O., Lar-
and sponge choanocytes. Plos Biology, 17(4), e3000226. https://doi. tillot, N., Wörheide, G., & Pisani, D. (2017). Improved modeling of
org/10.1371/journal.pbio.3000226 compositional heterogeneity supports sponges as sister to all other
65. Laumer, C. E., Fernandez, R., Lemer, S., Combosch, D., Kocot, K. M., animals. Current Biology, 27(24), 3864–3870 e3864. https://doi.org10.
Riesgo, A., Andrade, S. C. S., Sterrer, W., Sørensen, M. V., & Giribet, G. 1016/j.cub.2017.11.008
(2019). Revisiting metazoan phylogeny with genomic sampling of all 80. Kapli, P., & Telford, M. J. (2020). Topology-dependent asymmetry
phyla. Proceedings of the Royal Society B: Biological Sciences, 286(1906), in systematic errors affects phylogenetic placement of Ctenophora
20190831. https://doi.org/10.1098/rspb.2019.0831 and Xenacoelomorpha. Science Advances, 6(50), eabc5162. https://doi.
66. Steinmetz, P. R., Kraus, J. E., Larroux, C., Hammel, J. U., Amon- org10.1126/sciadv.abc5162
Hassenzahl, A., Houliston, E., Wörheide, G., Nickel, M., Degnan, B. M., 81. Kapli, P., Flouri, T., & Telford, M. J. (2021). Systematic errors in phyloge-
& Technau, U. (2012). Independent evolution of striated muscles in netic trees. Current Biology, 31(2), R59–R64. https://doi.org10.1016/j.
cnidarians and bilaterians. Nature, 487(7406), 231–234. https://doi. cub.2020.11.043
org/10.1038/nature11180 82. Nielsen, C. (2019). Early animal evolution: A morphologist’s view.
67. Dayraud, C., Alie, A., Jager, M., Chang, P., Le Guyader, H., Manuel, M., Royal Society Open Science, 6(7), 190638. https://doi.org10.1098/rsos.
& Queinnec, E. (2012). Independent specialisation of myosin II par- 190638
alogues in muscle vs. non-muscle functions during early animal evo- 83. King, N., & Rokas, A. (2017). Embracing Uncertainty in Reconstructing
lution: a ctenophore perspective. Bmc Evolutionary Biology [Electronic Early Animal Evolution. Current Biology, 27(19), R1081–R1088. https:
Resource], 12, 107. https://doi.org10.1186/1471-2148-12-107 //doi.org10.1016/j.cub.2017.08.054
68. Gueneli, N., McKenna, A. M., Ohkouchi, N., Boreham, C. J., Beghin,
J., Javaux, E. J., & Brocks, J. J. (2018). 1.1-billion-year-old porphyrins
establish a marine ecosystem dominated by bacterial primary pro-
ducers. Proceedings of the National Academy of Sciences of the United How to cite this article: Schierwater, B., Osigus, H.-.J.,
States of America, 115(30), E6978–E6986. https://doi.org10.1073/
Bergmann, T., Blackstone, N. W., Hadrys, H., Hauslage, J.,
pnas.1803866115
69. Schierwater, B., Holland, P. W. H., Miller, D. J., Stadler, P. F., Wieg- Humbert, P. O., Kamm, K., Kvansakul, M., Wysocki, K., &
mann, B. M., Wörheide, G., Wray, G. A., & DeSalle, R. (2016). Never DeSalle, R. (2021). The enigmatic Placozoa part 1: Exploring
ending analysis of a century old evolutionary debate: “unringing” the evolutionary controversies and poor ecological knowledge.
urmetazoon bell. Frontiers in Ecology and Evolution, 4(5). https://doi.
BioEssays, e2100080.
org10.3389/fevo.2016.00005
70. Laumer, C. E., Gruber-Vodicka, H., Hadfield, M. G., Pearse, V. B., Riesgo, https://doi.org/10.1002/bies.202100080
A., Marioni, J. C., & Giribet, G. (2018). Support for a clade of Placozoa