Investigations on Reproductive Affinities in Red Algae

Abstract

Allopatric speciation is the most widely accepted model proposed for speciation: once a population is divided by extrinsic
barriers, genetic flow is interrupted between these disjunct subpopulations, and reproductive isolation is established as
a by-product of the accumulation of genetic changes in these isolated populations. According to the biological species concept
(Mayr, 1942), the evolution of reproductive isolation is a defining characteristic of speciation, and reproductive isolation
contributes to the diversification of species by creating genetically independent lineages. It had been generally thought
that isolation is more difficult in marine populations, with frequent gene flow over large distances (Hoffmann, 1987; van
den Hoek, 1987; Norton, 1992; Shanks et al., 2003), as there seem to be far fewer extrinsic barriers in marine environments
than in terrestrial ones (Palumbi, 1994). Such attributes are considered to limit the isolation of a species into allopatric
populations less frequently, making allopatric speciation rarer (Mayr, 1954). However, recent molecular analyses have revealed
great genetic divergences among/within populations in various marine organisms, including macroalgae, and thus, the generalization
that speciation must be rare in marine habitats appears to be incorrect.

Full text

J. Seckbach and D.J. Chapman (eds.), Red Algae in the Genomic Age,
Cellular Origin, Life in Extreme Habitats and Astrobiology 13, 77–109
DOI 10.1007/978-90-481-3795-4_5, © Springer Science+Business Media B.V. 2010
Biodata of Mitsunobu Kamiya and John A. West, authors of “Investigations
on Reproductive Affinities in Red Algae”
Associate Professor Mitsunobu Kamiya, currently belongs to the Faculty of Marine
Bioscience, Fukui Prefectural University, Japan, and obtained his Ph.D. from the
University of Tsukuba in 1995. His research interests are in the areas of speciation
of red algae, ecophysiology of euryhaline algae, and algal allelopathy.
E-mail: mkamiya@fpu.ac.jp
Professor John A. West is currently a Professorial Fellow at the School of Botany,
University of Melbourne, Australia (1994–2009). He obtained his Ph.D. in
1966 from the University of Washington. Dr. West was a Professor of Botany,
University of California, Berkeley (1966–1994). His primary research interest is
on the biology of marine red algae.
E-mail: jwest@unimelb.edu.au
Mitsunobu Kamiya John A. West
77

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
MITSUNOBU KAMIYA1 AND JOHN A. WEST2
1Department of Marine Bioscience, Fukui Prefectural University,
Gakuencho, Obama, Fukui 917-0003 Japan
2School of Botany, University of Melbourne, Parkville, VIC 3010,
Australia
1. Introduction
Allopatric speciation is the most widely accepted model proposed for speciation: once
a population is divided by extrinsic barriers, genetic flow is interrupted between these
disjunct subpopulations, and reproductive isolation is established as a by-product of
the accumulation of genetic changes in these isolated populations. According to the
biological species concept (Mayr, 1942), the evolution of reproductive isolation is a
defining characteristic of speciation, and reproductive isolation contributes to the
diversification of species by creating genetically independent lineages. It had been
generally thought that isolation is more difficult in marine populations, with fre-
quent gene flow over large distances (Hoffmann, 1987; van den Hoek, 1987; Norton,
1992; Shanks et al., 2003), as there seem to be far fewer extrinsic barriers in marine
environments than in terrestrial ones (Palumbi, 1994). Such attributes are considered
to limit the isolation of a species into allopatric populations less frequently, making
allopatric speciation rarer (Mayr, 1954). However, recent molecular analyses have
revealed great genetic divergences among/within populations in various marine
organisms, including macroalgae, and thus, the generalization that speciation must
be rare in marine habitats appears to be incorrect.
Since the mid-twentieth century, crossing studies have been carried out in
various macroalgal groups to supplement morphological and cytological data, and
these results have contributed to delineation of species boundaries, inference of
evolutionary relationships, and disclosure of intraspecific diversities (cf. Mathieson
et al., 1981; Guiry, 1992; Brodie and Zuccarello, 2006). However, in recent years,
these processes are more effectively performed using molecular phylogenetic analyses
that were developed in the last 20 years, and, in contrast, crossing studies are
performed on only limited taxa. Some biologists consider that such crossing inves-
tigations provide more restricted information for taxonomic and phylogenetic
studies, and because of the time-consuming efforts required, they are forced to
forsake these “less fruitful” endeavors to concentrate on the molecular data in
studying macroalgal speciation. However, it is highly debatable that information
about reproductive affinities has little use and does not contribute to our under-
standing of algal evolution anymore. Can we fully explain the patterns and processes
79

MITSUNOBU KAMIYA AND JOHN A. WEST
of macroalgal speciation with only molecular evidence? Are not the reproductive
affinities helpful except for macroalgal taxonomy? To answer these questions, it is
necessary to demonstrate how the crossability information helps our understanding
of macroalgal speciation and will contribute to further investigations on the
genetic mechanisms of algal evolution.
The number of known red algal species (ca. 6,000 species) is much greater
than green (ca. 1,100 species) and brown algae (ca. 1,500 species) (Graham and
Wilcox, 2000). It is worthwhile to clarify how red algae diversify in their habitats
and how they adapt to the circumstances under natural selection. Red algae,
especially florideophycean algae, show a life history rather different from other
macroalgae. Nonflagellate spermatia from male gametophytes attach to receptive
trichogynes, hair-like cytoplasmic extensions of the female egg (carpogonium)
(Fig. 1), and the zygote grows into a microscopic diploid structure on the female
thallus. This diploid tissue, carposporophyte, is embedded in the parental female
thallus, and the combination of the carposporophyte tissue and the surrounding
female gametophytic tissue (pericarp) is called cystocarp (Fig. 2). Carpospores
released from the carposporophyte grow into diploid sporophytes (tetrasporo-
phytes), and their meiospores (tetraspores) develop into haploid gametophytes.
This unique triphasic life history is thought to enhance reproductive fecundity
and serve as an evolutionary compensation for loss of flagella (Searles, 1980;
Brawley and Johnson, 1992).
Reproductive affinities have been investigated in the widespread taxa of red
algae, because of the following advantages for crossing experiments. First, many
Figure 1. Female reproductive structure of Caloglossa leprieurii. Nonflagellate spermatia (S) from a
male gametophyte attached to a trichogyne (T), hair-like cytoplasmic extension of the female egg,
carpogonium (C).
80

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
algae require certain biotic and/or abiotic factors for gametogenesis and mating.
For example, Closterium and Volvox need the presence of the opposite mating
type to induce gametogenesis (Starr and Jaenicke, 1974; Tsuchikane et al., 2003).
Gamete interaction requires a certain minimal gamete concentration (Nečas,
1981; Togashi and Cox, 2004), and a certain similar proportion of the two gam-
ete types may be necessary for successful zygote formation (Richards and
Sommerfeld, 1974). Synchronous gamete release is also crucial for successful
fertilization of flagellated male and female gametes (see Santelices, 2002). In
contrast, red algal gametogenesis is relatively frequent and successive at least in
laboratory cultures, and hence, artificial hybridization can be carried out by
putting fertile male and female thalli together, without any induction for game-
togenesis and mating (e.g., Kamiya et al., 1997). Second, parthenogenetic devel-
opment of gametes, which is common in green and brown algae (De Wreede and
Klinger, 1988), is rare in red algae, except for apomictic entities of Ahnfeltiopsis
and Mastocarpus (Polanshek and West, 1977; West et al., 1978; Guiry and West,
1983; Masuda et al., 1984, 1987); hence, it is easy to infer fertilization through
the production of carposporophytes.
As the taxonomic implications of reproductive compatibility were well
documented by Mathieson et al. (1981) and Guiry (1992), in this chapter, we
will focus on the reproductive affinities, to provide useful information about the
pattern and process of red algal speciation. We will first document the variations
Figure 2-4. Various cystocarps produced in the artificial crosses of Caloglossa leprieurii. 2. Normal
cystocarp containing many carpsporangia inside. 3. Abortive cystocarp that failed to discharge
corpospores. 4. Pseudocystocarp with few carposporophyte tissues inside.
81

MITSUNOBU KAMIYA AND JOHN A. WEST
of reproductive fertility in intra- or interspecific crosses, and then elucidate how
the degree reproductive affinity is coupled with geographic distances, physiolog-
ical differences, and/or genetic variations. Lastly, we will discuss the perspective
and importance in assessing reproductive affinity for future research into algal
speciation.
2. Prezygotic Isolating Mechanism
Reproductive isolation is achieved in a variety of ways and falls into two broad
categories: prezygotic isolating mechanisms that act to prevent the formation of
hybrid zygotes, and postzygotic isolating mechanisms that bring about reproductive
failure at various stages after fertilization. In marine organisms, the simplest and
probably the most common prezygotic mechanism is spatial separation that pre-
vents genetic exchange between potentially interbreeding groups (see below for
detail). Even if potentially interbreeding groups are not geographically separated,
reproductive isolation can be maintained by ecological, temporal, or biochemical
differentiation between them. Some of these prezygotic isolating mechanisms may
operate simultaneously, and can make reproductive isolation more effective. As red
algae are the only group of macroalgae completely lacking the flagellated stages,
their fertilization is a random collision of two gametes, and as a result, some
spermatia may end up on the trichogynes of incompatible females. Therefore,
prezygotic isolating mechanisms are important to avoid a potential loss of fitness
producing abortive hybrid progenies.
2.1. ECOLOGICAL ISOLATION
In ecological isolation, even if potentially interbreeding groups live in the same
geographical area, they occupy different habitats and thus do not come into con-
tact with each other. However, the effect of this isolation mechanism is considered
to be limited in macroalgae, because free-living gametes are possibly exchanged
between the habitats. Richerd et al. (1993) performed crosses between Gracilaria
verrucosa (currently G. gracilis) collected from high and low tidal level and dem-
onstrated that the fertility between the different tidal levels was not significantly
different from that within the same tidal level. Recently, Engel et al. (2004) com-
pared the population structure of G. gracilis in light of the spatial positions of
populations with respect to height on the shore by analyzing seven microsatellite
loci. They indicated that high-shore individuals were more frequently restricted
to their original intertidal pools than low-shore individuals, and that height on
the shore significantly influenced migration among pools. High-shore populations
were found to experience longer periods of isolation from other intertidal pools at
the same shore, and as a result, low-shore populations were found to be more open
82

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
to migrants (Engel and Destombe, 2002; Engel et al., 2004). Such asymmetrical
gene flow between high- and low-shore populations can advance adaptation to
local ecological conditions. As the environmental conditions, such as light quality
and quantity, temperature, exposure time to air, and/or degree of wave exposure,
differ with coastal depth, the difference in growth depth may possibly develop into
ecological isolation. The Baltic Sea and adjacent regions provide a good example
of how salinity gradient is effective in interrupting gene flow. The salinity gradient
is created by a large inflow of freshwater into the Baltic Sea from over 200 rivers,
in combination with the semi-enclosed geographical position of the Baltic basin
(Bergström and Carlsson, 1994). In addition, this salinity gradient is stable year-
round owing to the long water turnover times. Despite the Baltic Sea flora having
developed from postglacial immigrants (8,000–4,000 BP), recent researches indi-
cate great reproductive and genetic diversities of several macroalgae within this
region (see Johannesson and André, 2006).
For example, Ceramium tenuicorne in the Baltic Sea and adjacent regions
has been well investigated based on reproductive crossability, salinity tolerance,
and genetic diversity. Many scientific names had been assigned to this alga
owing to its morphological variability, and Gabrielsen et al. (2003) suggested
that the specimens from the Skagerrak–Baltic region belong to the same species,
owing to their interfertility and almost identical sequences of the nuclear ribos-
omal internal transcribed spacer 2 (ITS2) and the plastid Rubisco spacer region.
However, the mitochondrial cox2-3 spacer analyses revealed five haplotypes in
C. tenuicorne from the Skagerrak–Baltic region; one of them was restricted in
the highest salinity site (Oslofjorden) where no other haplotype was detected,
and another occupied the low (Baltic) and medium (Kattegat) salinity sites
(Gabrielsen et al., 2002). In contrast to the cox2-3 spacer results, the genetic
analyses of random amplified polymorphic DNAs (RAPDs) suggested a con-
tinuous cline corresponding to the salinity gradient (Gabrielsen et al., 2002).
The authors imply that this incongruence of the two molecular data sets is pos-
sibly associated with the difference in the nuclear and mitochondrial inherit-
ance, and also with the vicariant event during the glacial period. High genetic
diversity has also been reported in other macroalgae (van Oppen et al., 1995b;
Valatka et al., 2000; Coyer et al., 2003; Johansson et al., 2003) as well as sea-
grass (Reusch et al., 2000; Reusch, 2002; Olsen et al., 2004) distributed in the
Baltic region. Although many Baltic macroalgae are considered to have been
introduced from the North Atlantic after the last glacial maximum, populations
inhabiting the Baltic have been influenced by different evolutionary forces
including severe bottlenecks, genetic isolation, and strong selection in this novel
habitat (Johannesson and André, 2006).
Physiological differentiation is observed in Ceramium tenuicorne from the
Skagerrak–Baltic region: the isolates from low and high salinity sites showed dif-
ferent growth responses in different salinities, which corresponded to the salinity
regimes of their original habitats, and their hybrid indicated an intermediate
83

MITSUNOBU KAMIYA AND JOHN A. WEST
pattern relative to that of the parents (Rueness, 1978; Rueness and Kornfeldt,
1992; Bergström and Kautsky, 2006). Bergström et al. (2003) observed a strong
reduction in sexual reproduction and an increased tetrasporophyte dominance
along the salinity gradient in C. tenuicorne from its inner distribution limit in the
Baltic Sea. Although it is unknown whether this reproductive strategy is geneti-
cally fixed in these entities, it is presumed to affect the genetic exchange along the
Baltic coasts.
A similar genetic disjunction between the eastern North Atlantic and
Baltic Sea coasts was observed in Phycodrys rubens. Two genetic groups were
recognized around these regions based on allozymes and RAPDs data: each
genetic group occupied the European outer coasts and the North Sea/Baltic
coasts, respectively, and these two genetic groups co-occurred in the Skagerrak
and Kattegat regions (van Oppen et al., 1995a, b). As salinities decreased below
20 psu, the relative growth rate of marine strains of P. rubens from the North
Sea was severely reduced when compared with the brackish strains from the
Baltic Sea (Rietema, 1991). Ecotypic differentiations in salinity response have
been observed in many other macroalgae distributed around this region
(Russell, 1985, 1994; Thomas et al., 1990; Bäck et al., 1992; Rietema, 1993,
1995; Kristiansen et al., 1994; Serrão et al., 1996), and it is highly possible that
this salinity gradient could be responsible for generating adaptive differentia-
tion as well as may act as a barrier for gene flow among these ecotypes
(Middelboe et al., 1997).
Difference in upper or lower temperature tolerance is also an important
factor for algal ecological isolation. In the case of Chondrus crispus from two
sites of Atlantic Canada, the strain from Bay of Fundy indicated a faster growth
rate at 10–20°C than the strain from Gulf of St. Lawrence (Chen and Taylor,
1980). Cystocarps were produced on the female from the Gulf of St. Lawrence
in the presence of the male from the Bay of Fundy, whereas no reaction was seen
in the reciprocal cross. Their different temperature response may maintain “habi-
tat segregation,” and as a result, may restrict the genetic exchange between these
localities, though Chopin et al. (1996) suggested low genetic diversities of
Chondrus crispus around the Atlantic Canada. The difference in the upper or
lower temperature for maturation within the same species, which have been
reported in many kinds of red algae (e.g., Molenaar and Breeman, 1994;
Molenaar et al., 1996; West et al., 1996), may also be an important factor for
ecological isolation.
2.2. TEMPORAL ISOLATION
Temporal isolation is similar to ecological isolation, but with the separation
occurring in time rather than space; the entities reproduce in different seasons or
even different times of the day. Currently, there is one good instance for temporal
84

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
isolation of red algae. The western Pacific strains of Chondrus ocellatus show dif-
ferent photoperiodic response: the strains from Korea, China, and some Japanese
sites form gametangia under long-day conditions, whereas those from two other
Japanese sites (Enoshima and Sumoto) require short-day conditions for gameto-
genesis (Brodie et al., 1993). The photoperiodically different strains are interfertile
once they produce gametes under their appropriate condition, and the responses
to daylength in their F1 gametophytes suggest Mendelian inheritance of this
photoperiodic property.
2.3. BIOCHEMICAL ISOLATION
On biochemical isolation, it was found that the entities failed to fertilize success-
fully because of phenotypic incompatibilities. In the case of red algae, sexual
reproduction is brought about by the attachment of spermatia to trichogynes.
After the localized cell-wall breakdown of the trichogyne, the spermatial nuclei
are transported along the trichogyne into the carpogonium, where karyogamy
takes place (Mine and Tatewaki, 1994; Pickett-Heaps and West, 1998). Magruder
(1984) demonstrated that spermatia from Aglaothamnion cordatum (as A. neglectum)
specifically bind with trichogynes and hairs of female thalli, and that fimbriate
cone-shaped appendages projecting from each end of the spermatium are respon-
sible for the initial binding with trichogynes. The lectin-/carbohydrates-binding
experiments of Ceramiacean species suggested that the attachment of spermatia
to trichogynes is mediated by particular carbohydrates on spermatial surface
and complementary carbohydrates-binding receptors on trichogynes (Kim and
Fritz, 1993; Kim et al., 1996; Kim and Kim, 1999). In Antithamnion sparsum,
d-mannose and l-fucose were involved in gamete recognition, whereas the latter
was not involved in Aglaothamnion oosumiense (Kim et al., 1996; Kim and Kim,
1999). Although these glycoproteins must be important for fertilization, which
can be blocked by adding the complimentary sugars or lectins, it is still uncer-
tain whether these glycoproteins are responsible for species recognition. Quite
recently, Kim et al. (2008) undertook a comparison of the proteome among
eight isolates of the Bostrychia radicans/moritziana species complex showing
various reproductive reactions with each other. The male and female isolates
had 3.7–7.1% sex-specific proteins, and the lack of any shared sex-specific pro-
teins across all isolates may suggest rapid evolution of these proteins (Kim et al.,
2008). Such a proteome analysis is expected to become an important cue for the
elucidation of the molecular mechanisms of sex as well as species recognition
in red algae.
In Chlamydomonas gametes, sex-cell contact at fertilization is based on the
complementarity between special mating-type glycoproteins (agglutinin) located
on the flagellar membranes (Goodenough and Adair, 1989), and the gene
sequences from these proteins are strikingly different between the closely related
85

MITSUNOBU KAMIYA AND JOHN A. WEST
species (Lee et al., 2007). Bolwell et al. (1980) reported that membrane fractions
isolated from either eggs or sperms of Fucus serratus inhibited fertilization in a
species-specific manner, and suggested that this inhibitory activity was associated
with a high-molecular-weight glycoprotein containing a-fucose and a-mannose.
The species-specificity of such a glycoprotein was confirmed by immunological
studies (Jones et al., 1988, 1990).
By considering the fact that spermatial binding to trichogynes is not species-
specific, but occurs between closely related species (Magruder, 1984), the bio-
chemical or mechanical isolation, if present, can be presumed to work after the
attachment of spermatia to trichogyne. Engel et al. (1999) evaluated the success
rate of male fertilization in a natural population of Gracilaria gracilis by assessing
the individual contribution of different males to carposporophytes, and by deter-
mining the paternity using two microsatellite loci. They found significant inter-
male differences in success rate of fertilization, regardless of the distance between
the male and female gametophytes, and suggested the possibility of nonrandom
mating, resulting either from female choice or from male-to-male competition.
This nonrandom mating was confirmed by crossing experiments using multiple
individuals as sources of spermatia (Engel et al., 2002), and these results may
implicate that inequality of male performance in postadhesion events generates
nonrandom mating.
The behavior of spermatial nuclei after attachment of spermatia to tricho-
gynes has been observed using time-lapse video microcopy. In Rhodomelaceae,
the spermatial nucleus divides once after the attachment and the two nuclei are
injected into the trichogyne, one moving down to the base and the other moving
up to the tip (Pickett-Heaps and West, 1998; Wilson et al., 2002, 2003). Pickett-
Heaps and West (1998) observed the attachment of multiple spermatia to a
trichogyne and the movement of these nuclei within trichogynes in Bostrychia
moritziana. They suggested that the transport systems of different gamete
nuclei may interact with one another during the active transfer toward the car-
pogonial nucleus. Although there is still much to learn about gamete recogni-
tion and isolation mechanism before examining karyogamy in red algae,
continued research, using methodologies as cited earlier may provide better
understanding regarding what and how prezygotic biochemical isolating mech-
anisms function in red algae.
3. Postzygotic Isolating Mechanism
Although no prezygotic isolating mechanism has been established and zygotes
are formed between two populations, genetic differences have become very
significant, such that the resulting hybrids are less viable or less fertile than
the parents. Artificial crossing experiments, usually performed among closely
related species or within the same species, make it possible to reveal potential
86

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
compatibility and the degree of reproductive isolation between them. In the case
of red algae, various kinds of postzygotic isolating phenomena, such as the for-
mation of sterile/inviable F1 gametophytes or sporophytes, abortive cystocarps
(Fig. 3), or pseudocystocarps (Fig. 4), have been frequently observed using cul-
ture strains.
3.1. INVIABLE OR STERILE F1 GAMETOPHYTES
Although production of fertile F1 tetrasporophytes is sometimes regarded as a
successful cross, it is possible that their tetraspore germlings do not grow well or
grow into sterile F1 gametophytes. Production of such inviable or sterile F1 game-
tophytes is evident as the first stage of postzygotic isolation. Miura et al. (1992)
performed hybridization using pigmentation variants of Porphyra tenera and
P. yezoensis, which were compatible and produced fertile F1 heterozygous sporo-
phytes showing wild-type color. However, most of the F1 gametophytic germlings
died at the four-cell stage, and very few single-colored or chimeric gametophytes
survived. Meiosis was observed to occur during conchospore germination in
P. yezoensis, based on the evidences of chimeric blade formation (Ohme and
Miura, 1988) and cytogenetic analyses (Ma and Miura, 1984; Shimizu et al.,
2008; but see Wang et al., 2006 for different position of meiosis), and thus, the
breakdown of F1 gametophytes could have probably been caused by defective
meiosis (Miura et al., 1992).
Kudo and Masuda (1986) demonstrated a variety of reproductive reactions
between crosses of Polysiphonia akkeshiensis and P. japonica from central and
northern Japan. Polysiphonia akkeshiensis females were intersterile with P. japonica
males, while the reciprocal crosses frequently produced fertile F1 tetrasporophytes.
However, the tetrasporelings did not grow well in some crosses, or in other
crosses, they grew into sterile gametophytes. Rueness (1973) obtained fertile F1
tetrasporophytes in the crosses between Polysiphonia boldii from Texas, USA, and
P. hemisphaerica from Scandinavia. Although more than 1,000 tetraspores were
discharged from the hybrid tetrasporophytes, most of them failed to develop after
a few divisions and only a few spores grew into fertile gametophytes. Although F1
tetrasporophytes released tetraspores in most of the crossings of Digenea simplex
from the Atlantic and Caribbean Seas, the percentage of healthy tetrasporelings
were apparently higher in the crossings between the adjacent populations
(77–100%) than in the crossings between the distant populations (3–43%) (Pakker
et al., 1996).
Meiosis has been observed in the formation of tetraspores in some flo-
rideophycean species, such as Wrangelia plumosa and Antithamnionella pacifica
(Goff and Coleman, 1990), and some parasitic species of Janczewskia,
Levringiella, Gonimophyllum (Kugrens and West, 1972), and Choreocolax (Goff
and Coleman, 1984). However, Scagelia pylaisaei has been observed to undergo
87

MITSUNOBU KAMIYA AND JOHN A. WEST
meiosis during tetraspore germination like Porphyra (Goff and Coleman, 1990).
According to the cytological observations of Palmaria palmata from Atlantic
Canada and Ireland, which produced inviable F1 gametophytes, the hybrid
tetraspo rophytes showed complex chromosome pairings, which may explain
meiotic abnormality (van der Meer, 1986, 1987). Deficient meiosis may probably
be one of the reasons for inviable hybrid gametophytes, but cannot easily explain
various abnormalities, from no germination of tetraspores to production of abor-
tive cystocarps on F1 hybrid females.
3.2. ABORTIVE CYSTOCARPS OR INVIABLE F1 SPOROPHYTES
In contrast to most of the organisms in which a zygote is released from the game-
tophyte to produce a diploid individual, a red algal zygote grows into a multicel-
lular carposporophyte and discharges diploid carpospores while attached to the
female gametophyte, and hence, cystocarp development is a good indicator of
reproductive affinity.
Abortive cystocarps (Fig. 3) that fail to discharge carpospores or are dis-
rupted before sporulation, or inviable F1 sporophytes, have also been observed in
various crosses. For instance, abortive cystocarps were produced between a
Polysiphonia acuminata male from California and a P. japonica female from
Korea, whereas nonviable F1 tetrasporophytes appeared in the reciprocal cross
(Yoon and West, 1990). The cross between a male Gracilaria foliifera from the UK
and a G. sp. female from Italy produced cystocarps that failed to discharge carpo-
spores, while no reaction occurred in the reciprocal cross (Bird and McLachlan,
1982). Formation of abortive cystocarps or nonviable F1 sporophytes undoubtedly
reduces fitness, and such partially compatible pairs are usually found from
geographically distant populations (Table 1).
Developmental patterns of the carposporophyte are diverse within the
florideophycean algae and have been used as key systematic characters for
generic or higher taxonomic ranks (Hommersand and Fredericq, 1990), whereas
the physiological or genetic studies on this stage have not progressed. It is
known that carposporophytes of some red algae discharge many carpospores
for several weeks, with the total numbers of spores released reaching into the
thousands (Boney, 1960; Wilce and Sears, 1991; West and McBride, 1999;
Kamiya and Kawai, 2002). The number of cystocarps and discharged carpo-
spores per cystocarp is apparently various among the cross pairs even in the
same species. More cystocarps are produced and more carpospores are dis-
charged from each cystocarp in the self-crossing than the outcrossing between
the distantly distributed gametophytes (M. Kamiya and J. A. West, unpublished
data, 2008). A negative correlation between the number of cystocarps and the
geographical distance has been reported by Zuccarello and West (1995). Such
variations of carposporophyte productivity possibly represent the intraspecific
88

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
difference of reproductive affinity, and more detailed analyses on carposporo-
phyte development and carpospore formation are required for elucidating the
genetic impact of outcrossing.
3.3. PSEUDOCYSTOCARPS
A cystocarp-like swelling, usually called pseudocystocarp (Fig. 4), is frequently
observed in red algal crosses. Although this structure develops pericarps and
an ostiole, there is no or little initial carposporophyte tissue formed and no
carpospores are discharged. McLachlan et al. (1977) observed pseudocystocarp
formation in the crosses between Gracilaria foliifera from the UK and G. tikva-
hiae (as G. sp.) from Nova Scotia. Pseudocystocarps were also produced in the
cross between a G. cervicornis male and a G. mammillaris female from Brazil,
but no pseudocystocarps occurred in the reciprocal cross (Plastino and Oliveira,
1988). Most of the cross pairs forming pseudocystocarps were isolated from
geographically distant populations, between which genetic exchange must be
quite low (Table 1).
Pseudocystocarps are formed in crosses showing various degrees of genetic
diversity; the sequence divergence of the Rubisco spacer and its flanking regions
is found to be 0.6–1.7% in Bostrychia radicans (Zuccarello and West, 1997),
0.4–2.6% in Caloglossa leprieurii (Kamiya et al., 1998), 3.7% in C. postiae
(Kamiya et al., 1999), and 0.6–5.8% in C. monosticha (M. Kamiya and J. A.
West, unpublished data, 2008). Crossability, including production of pseudo-
cystocarps, can be considered as a symplesiomorphy, a maintained ancestral
characteristic, and hence, the interfertile entities, regardless of whether fully or
partially, may not necessarily be close phylogenetically (Donoghue, 1985). In
fact, there are some reports of the inconsistency between crossability and genetic
distance (e.g., Pakker et al., 1996; Zuccarello and West, 1997), and hence, cau-
tion must be used in assessing reproductive compatibility as a phylogenetic simi-
larity indicator.
The mechanism causing the improper development of carposoporphyte
(i.e., pseudocystocarp) is poorly known. McLachlan et al. (1977) reported that
syngamy occurred in these crosses as pseudocystocarps did not form either in
the presence of heat-killed spermatia or filtrates from suspensions of viable
spermatia. Destombe et al. (1990) indicated that the frequency of pseudocysto-
carp production increased with the age of the male gametes. Boo and Lee
(1983) observed the early discontinuance of carposporophyte development in a
cross of Antithamnion defectum male × A. sparsum female. Pseudocystocarp
formation is assumed to be under the control of interactions between maternal
haploid tissue and zygotic diploid tissue (Hommersand and Fredericq, 1990).
As pseudocystocarps are usually produced between morphologically similar or
identical entities, the formation of such structures has been considered as
89

MITSUNOBU KAMIYA AND JOHN A. WEST
Table 1. A summary of the red algal crossing results.
Species (distribution) Compatibility Comments Reference
Bangiales
Porphyra anagusta (Tokyo,
JPN), P. pseudolinearis
(Miyagi, JPN), P. tenera
(Fukushima, Miyagi and
Tokyo, JPN), P. umbilicalis
(Tokyo, JPN) and
P. yezoensis (Tokyo, JPN)
F1 gametophytes fertile in the
crosses among P. anagusta,
P. pseudolinearis, and
P. umbilicalis. These species
are fertile with P. tenera and
P. yezoensis, but most of the F1
gametophytes are abnormal.
Suto (1963)
Porphyra tenera (green
mutant; locality unknown)
and P. yezoensis (wildtype,
green and red mutants;
JPN)
F1 sporophytes fertile, but
most F1 gametophytes become
extinct at the four-cell stage in
either interspecific or
intermutant crosses.
F1 gametophytes
between the
different color
mutants were
chimeric.
Miura et al.
(1992)
Palmariales
Palmaria palmata var.
palmata (Canada and
Ireland) and var. sobolifera
(Ireland)
Viable F1 gametophytes
produced in the intervariety
crossings of the Irish strains,
but inviable F1 gametophytes
produced between these Irish
strains and Canadian var.
palmata.
van der Meer
(1987)
Gracilariales
Gracilaria foliifera (UK), G.
tikvahiae (Atlantic Canada),
G. sp.1 (Italy) and G. sp.2
(Pacific USA)
Fertile cystocarps failed to
discharge spores between
G. foliifera male and G.
sp.1 female. No reaction or
pseudocystocarps produced in
other crosses.
McLachlan
et al. (1977),
Bird and
McLachlan
(1982)
Gracilaria multipartita
(France, Ireland and UK)
and G. tikvahiae (Pacific
Canada, North Carolina
and Texas in USA)
Fertile F1 tetrasporophytes
produced in the intraspecific
crosses, but no reaction in the
interspecific crosses
The strain from
North Carolina
was more similar
to the European
strains than the
Texas strain.
Guiry and
Freamhainn
(1986)
Gracilaria verrucosa (now a
synonym of G. gracilis;
Pacific Canada and UK)
No reproductive reaction The chromosome
number was
different between
them (n = 32 for
British strain and
n = 24 for
Canadian strain).
Bird et al.
(1982)
Gracilaria verrucosa (now a
synonym of G. gracilis;
Shinori and Kikonai, JPN)
and G. vermiculophylla
(Akkeshi, JPN)
F1 gametophytes fertile in all
the crosses.
Yamamoto
and Sasaki
(1987, 1988)
(continued)
90

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
Table 1. (continued)
Species (distribution) Compatibility Comments Reference
Gracilaria verrucosa (now a
synonym of G. gracilis;
Argentina, Pacific Canada,
France, Norway and UK)
Viable or inviable F1 tetraspo-
rophytes produced among the
strains from Argentina, France,
Norway, and UK. The strain
from Pacific Canada interster-
ile with that from UK.
Rice and Bird
(1990)
Gracilaria verrucosa (now a
synonym of G. gracilis; five
sites in France)
Both normal and abortive
cystocarps produced in the
most crosses attempted.
Fertility was
significantly
different among the
females, but not
different among
the males.
Richerd et al.
(1993)
Gracilaria cervicornis (Brazil
and Pacific USA),
G. mammillaris (Brazil) and
G. aff. verrucosa (now a
synonym of G. gracilis;
Brazil and Pacific USA)
Normal cyctocarps produced
in Brazilian male × US female
of G. cervicornis but no
reaction in the reciprocal cross.
Normal cyctocarps produced
between the strains of G. aff.
verrucosa. Pseudocystocarps
produced in the Brazilian male
of G. cervicornis ×
G. mammillaris female, but no
reaction in the reciprocal cross.
Plastino and
Oliveira (1988)
Gracilaria tikvahiae
(Canada and Florida)
Incomplete. van der Meer
(1986)
Gelidiales
Gelidium pulchellum (France
and Ireland), G. pusillum
(France, Ireland, Norway
and UK)
Fertile F1 tetrasporophytes
produced in the intraspecific
crosses, but no reaction in the
interspecific crosses.
Fredriksen
et al. (1994)
Gigartinales
Gigartina teedii (now a
synonym of Chondracanthus
teedei; Brazil, France,
Greece, Ireland, Sicily and
UK)
F1 tetrasporophytes fertile. Growth rate was
different between
the Mediterranean
and Atlantic
strains. Gross
morphology and
color were different
among these
strains.
Guiry (1984),
Guiry et al.
(1987)
Petrocelis middendorfii
(Alaska) and P. franciscana
(now a synonym of
Mastocarpus papillatus; two
sites in California)
Carpospores produced. Polanshek and
West (1975)
(continued)
91

MITSUNOBU KAMIYA AND JOHN A. WEST
Table 1. (continued)
Species (distribution) Compatibility Comments Reference
Male of Gigartina papillata
(now a synonym of
Mastocarpus papillatus;
seven sites in California)
and female of Petrocelis
middendorfii (eight sites in
California)
Carpospores produced in some
crosses, but not in other
crosses.
Polanshek and
West (1977)
Gigartina pacifica (now a
synonym of Mastocarpus
pacificus; four sites in
Hokkaido, JPN)
F1 sporophytes produced in the
most crosses.
Masuda et al.
(1984)
Ahnfeltiopsis concinna
(Hawaii and six sites in
JPN)
Fertile F1 gametophytes pro-
duced.
No morphological
difference among
them.
Masuda and
Kogame (1998)
Mastocarpus sp. (now
regarded as M. yendoi; eight
sites in JPN)
Fertile F1 sporophytes pro-
duced in the most crosses.
Masuda et al.
(1987)
Gigartina stellata and
Petrocelis cruenta (now syn-
onyms of Mastocarpus stel-
latus; France, Iceland,
Ireland, Portugal, Spain
and UK)
Carpospores released among
the northern populations or
among the southern
populations, but no reaction
between them.
The northern and
southern
populations were
morphologically
distinguishable.
Guiry and
West (1983)
Gigartina agardhii (now a
synonym of Mastocarpus
jardinii; seven sites in
California)
Carpospores produced and
germinated in the most crosses
attempted.
West et al.
(1978)
Chondrus crispus (Atlantic
Canada, France, Germany,
Iceland, Norway, two sites in
Spain, five sites in Ireland,
and four sites in UK)
Carpospores released in all the
crosses attempted, and fertile
tetrasporophytes produced in
10% of the crosses
Guiry (1992)
Chondrus crispus (Bay of
Fundy and Gulf of St.
Lawrence, Nova Scotia,
Canada)
Cystocarps produced on the
female from Gulf of St.
Lawrence with the male from
Bay of Fundy, but no reaction
in the reciprocal cross.
The gross morphol-
ogy and develop-
mental response to
temperature were
different between
the two strains.
Chen and
Taylor (1980)
Chondrus pinnulatus f. pin-
nulatus (Abashiri,
Hanasaki, Muroran and
Oshoro, JPN), C. pinnulatus
f. armatus (now C. armatus;
Fukaura, Kikonai, Nemuro,
Oma and Oshoro, JPN)
F1 gametophytes fertile in the
intraformae crosses. Results of
interformae crosses variable
from no reaction to formation
of fertile F1 gametophytes.
Upper temperature
tolerance was dif-
ferent between the
two formae.
Brodie et al.
(1997)
(continued)
92

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
Table 1. (continued)
Species (distribution) Compatibility Comments Reference
Chondrus nipponicus (two
sites in JPN), C. ocellatus f.
crispoides (Aomori, JPN)
and f. ocellatus (China,
Korea and three sites in
JPN)
Fertile F1 tetrasporophytes pro-
duced in the intraformae
crosses. Results of interformae
crosses were variable from no
reaction to formation of viable
F1 gametophytes. Fertile F1 tet-
rasporophytes produced
between C. nipponicus and f.
crispoides.
In f. ocellatus, the
strains from China,
Korea, and one
Japanese site
required long-day
condition for game-
togenesis, whereas
the other two
Japanese strains
required short-day
condition.
Brodie et al.
(1993)
Digenea simplex (Atlantic
USA, Netherlands Antilles,
Cape Verde Islands and
Western Australia)
Various percentages of healthy
tetrasporelings produced in the
crossings, except for the
Western Australian strain,
which produced sterile F1 spo-
rophytes with the strains from
Atlantic USA and Cape Verde
Islands, but showed no reac-
tion with other strains.
Despite the reduced
level of sexual com-
patibility between
Caribbean and
Cape Verde Islands
isolates, they shared
position in the
RAPD analysis and
showed similar
temperature
responses.
Pakker et al.
(1996)
Ceramiales
Aglaothamnion byssoides
(Sweden and Atlantic USA)
and A. furcellariae (France)
(now synonyms of
A. tenuissimum)
The Swedish A. byssoides inter-
sterile with the American A.
byssoides, but produced F1 tet-
rasporophytes with A. furcel-
lariae.
These interfertile
strains were mor-
phologically similar
to each other.
L’Hardy-Halos
and Rueness
(1990)
Aglaothamnion byssoides
(now a synonym of A. ten-
uissimum; Sweden) and A.
tenuissimum var. mazoyerae
(Italy)
F1 tetrasporophytes produced
in the interspecific crosses.
Furnari et al.
(1998)
Antithamnion defectum
(Pacific USA) and A. spar-
sum (two sites in Korea)
Viable carpospores released
between A. sparsum male and
A. defectum female, but no
carpospores released in the
reciprocal cross.
Boo and Lee
(1983)
Antithamnion plumula var.
bebbii (now a synonym of
Pterothamnion crispum;
three sites in Norway and
Sweden), var. crispum (now
a synonym of P. crispum;
two sites in UK) and var.
plumura (four sites in
Norway and UK)
Gonimoblasts developed in the
intravariety crosses and
between all strains of var. beb-
bii and var. plumura from
Hoftoy in Norway and
Plymouth in UK. No reaction
in other intervariety crosses.
Interfertile strains
of var. bebbii and
var. plumura were
isolated from adja-
cent sites in
Norway.
Sundene
(1975)
(continued)
93

MITSUNOBU KAMIYA AND JOHN A. WEST
Table 1. (continued)
Species (distribution) Compatibility Comments Reference
Antithamnion plumula var.
bebbii (now a synonym of
Pterothamnion crispum;
Sweden) and var. plumura
(UK)
F2 gametophytes fertile in the
intervariety crosses.
The heterozygous
tetrasporophytes
always showed the
var. plumula pheno-
type and their tet-
raspores grew into
the two varieties in
equal numbers.
Rueness and
Rueness (1975)
Callithamnion byssoides
(now a synonym of
Aglaothamnion tenuissimum;
Norway and four Atlantic
sites in USA) and C. halliae
(now a synonym of A. hal-
liae; Texas, USA)
Callithamnion halliae fertile
with the four American strains
but sterile with the Norwegian
strain of C. byssoides. The four
American strains were interfer-
tile with each other, but sterile
with the Norwegian strain.
Thallus color was
different between
C. byssoides and
C. halliae but they
were morphologi-
cally indistinguish-
able. Their hybrids
showed
intermediate color.
Spencer et al.
(1981)
Callithamnion boergesenii
(now a synonym of
Aglaothamnion boergesenii;
Puerto Rico) and C.
byssoides (now a synonym
of A. tenuissimum; North
Carolina and Georgia,
USA)
The two American strains
of C. byssoides interfertile with
each other, but sterile with C.
boergesenii.
Aponte and
Ballantine
(1990)
Ceramium strictum (now a
synonym of C. tenuicorne;
Norway and Atlantic USA)
and C. tenuicorne (Baltic
Sea)
The US C. strictum intersterile
with other strains and F1 game-
tophytes produced between the
Norwegian C. strictum and
C. tenuicorne.
The morphology
and growth
response against
salinity regime were
similar between the
US and Norwegian
C. strictum, but
obviously different
between C. strictum
and C. tenuicorne.
Rueness (1978)
Ceramium strictum (now a
synonym of C. tenuicorne;
Denmark, Norway and
Sweden)
F1 gametophytes fertile. Salinity tolerance
was different among
the strains and their
hybrids showed
intermediate
responses between
the parents.
Rueness and
Kornfeldt
(1992)
Ceramium tenuicorne (three
strains from Skagerrak and
six strains from Baltic Sea)
Most intraspecific crosses
attempted successful, resulting
in germinating carpospores.
Gabrielsen
et al. (2003)
(continued)
94

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
Table 1. (continued)
Species (distribution) Compatibility Comments Reference
Ceramium aff. rubrum (now
a synonym of C. virgatum;
each two sites in
Washington, USA and
Nova Scotia, Canada)
F1 gametophytes fertile
between one American strain
and one Canadian strain, but
no cystocarps or no viable
carpospores produced in the
other crosses.
Garbary
(1988)
Caloglossa continua ssp.
axillaris (now a synonym of
C. monosticha; north
Australia), ssp. continua
(JPN, Taiwan and
Vietnam), C. monosticha
(Indonesia, Singapore and
west Australia ) and
C. saigonensis (Malaysia)
Fertile F1 gametophytes
produced within ssp. continua
and between the Australian
and Indonesian C. monosticha.
Variable reaction observed in
the most other crosses, from no
reaction to production of
inviable F1 gametophytes.
Kamiya et al.
(2003)
Caloglossa intermedia
(Georgia and South
Carolina, USA)
Fertile F1 gametophytes
produced.
Kamiya et al.
(2000)
Caloglossa leprieurii (South
Africa, Venezuela, Peru,
Atlantic USA and each two
sites in JPN and Singapore)
Five mating groups recognized,
and no reaction or only
pseudocystocarps observed
between the different mating
groups.
One mating
group was
morphologically
distinguishable
from the others.
Kamiya et al.
(1998),
Kamiya (2004)
Caloglossa leprieurii (now
regarded as C. vieillardii;
Fiji, three sites in JPN and
four sites in Australia)
Four mating groups recog-
nized. Inviable F1 gameto-
phytes or sporophytes
produced between the Japanese
and Western Australian mating
groups, while no reaction or
only pseudocystocarps
observed between the other
mating groups.
Kamiya et al.
(1995),
Kamiya (2004)
Caloglossa monosticha
(six sites in Australia and
three sites in Indonesia)
Pseudocystocarps, inviable F1
sporophytes or inviable F1
gametophytes produced
between the two Australian
strains and the other strains.
Fertile F1 gametophytes
produced in the most other
crosses.
Kamiya and
West (2008)
Caloglossa postiae
(Australia and two sites in
JPN)
Pseudocystocarps produced
between the Australian and
Japanese strains. Fertile F1
gametophytes
produced between the
Japanese strains.
Kamiya et al.
(1999)
(continued)
95

MITSUNOBU KAMIYA AND JOHN A. WEST
Table 1. (continued)
Species (distribution) Compatibility Comments Reference
Bostrychia radicans (seven
sites in Atlantic North
America and seven sites in
Pacific North America)
Viable F1 sporophytes
produced among the Pacific
populations. At least six
mating groups recognized
along the Atlantic coast, and
pseudocystocarps were
produced between some
mating groups. Inviable F1
gametophytes produced in the
cross between South Carolina
and Pacific Mexico.
There was a
negative correlation
between the
number of
cystocarps and
geographical
distance.
Zuccarello and
West (1995)
Bostrychia moritziana (New
Zealand, Fiji, Indonesia,
and several sites in Australia
and South Africa)
The Indonesian strain sterile
with any other strains. The
South African strain displayed
a lower compatibility with
those from Australia, New
Zealand and Fiji, which
produced viable F1 sporophytes
with each other.
Zuccarello
et al. (1999)
Bostrychia moritziana
(Australia and South
Africa) and B. radicans
(Australia, Pacific Mexico,
Peru, Brazil, Venezuela and
Atlantic USA)
Carpospores released in the
intraspecific cross of B. moritz-
iana, and at least four mating
groups recognized in B. radi-
cans. No reaction observed
between the different mating
groups or in the interspecific
crosses.
Zuccarello and
West (1997,
2003)
Laurencia japonensis (three
sites in JPN), L. nipponica
(three sites in JPN) and L.
okamurae (three sites in
JPN)
Fertile F1 gametophytes pro-
duced in all the intraspecific
crosses. Only pericarps devel-
oped in some crosses between
the L. okamurae male and L.
nipponica female, while no
reaction observed in the other
interspecific crosses.
Abe and
Masuda (1998)
Murrayella periclados
(Guam, French Polynesia,
Fiji, Indonesia, Mindanao
and Cebu in Philippines)
Carpospores germinated in
most crosses, but failed to ger-
minate between the male from
French Polynesia, Cebu or
Indonesia and the female from
Mindanao or Guam.
Zuccarello
et al. (2002)
Polysiphonia ferulacea (now
a synonym of Neosiphonia
ferulacea; North Carolina,
USA and Bermuda, UK)
No response observed between
the two strains.
Chromosome
number was
different between
the two strains.
Kapraun
(1977)
(continued)
96

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
indicative of an evolutionary link between them (Guiry et al., 1987). In the
absence of spermatia, formation of pseudocystocarps has been observed occa-
sionally in Bostrychia (J. A. West, unpublished data, 2008), Caloglossa (Tanaka
and Kamiya, 1993), and Gracilaria (Bird and McLachlan, 1982). Zuccarello
et al. (2002) tried intraspecific hybridization among worldwide populations of
Spyridia filamentosa and Murrayella periclados, and pseudocystocarps were
produced not only in unsuccessful crosses, but also in successful crosses that
produced normal cystocarps.
4. Relationships Between Reproductive Affinities, Geographic Distance,
and Genetic Diversity
Positive compatibility has been observed in many crossing experiments between
geographically distant populations, and in some cases, their genetic similarities were
also confirmed by DNA markers. For example, interfertility was demonstrated in
Chondrus crispus from the east and west Atlantic coasts (Guiry, 1981, 1992). In the
molecular phylogenetic analyses of C. crispus from Europe and Pacific Canada
Species (distribution) Compatibility Comments Reference
Polysiphonia acuminata
(Pacific USA) and P.
japonica (Korea)
Abortive cystocarps produced
between P. acuminata male and
P. japonica female and F1
tetrasporophytes inviable in the
reciprocal cross.
Yoon and West
(1990)
Polysiphonia akkeshiensis
(three sites in JPN) and P.
japonica (five sites in JPN)
(now regarded as
Neoshiphonia harveyi)
F2 tetrasporophytes fertile in
the intraspecific crosses.
Results of the interformae
crosses variable from no
reaction to formation of fertile
F2 tetrasporophytes.
The morphological
difference between
the two species was
not maintained
in the culture
conditions.
Kudo and
Masuda (1986)
Polysiphonia boldii (Texas,
USA) and P. hemisphaerica
(now a synonym of
P. boldii; Scandinavia)
Most F1 gametophytes inviable. Rueness (1973)
Spyridia filamentosa (South
Australia, Philippines,
Pacific Mexico and Puerto
Rico)
Carpospores released in the
cross between the Australian
male and Mexican female,
while no reaction or only pseu-
docystocarps produced in other
crosses.
Zuccarello
et al. (2002)
Table 1. (continued)
97

MITSUNOBU KAMIYA AND JOHN A. WEST
based on the ITS1 sequence data, the genetic distance was not to be correlated
with the geographic distance at all, and hence, multiple transatlantic dispersal was
suggested (Hu et al., 2007). Interfertility between geographically distant popu-
lations was also demonstrated in Bostrychia radicans from both sides of South
America (Pacific Mexico vs. Venezuela, or Peru vs. Brazil), and there were only
a few site changes at the Rubisco spacer plus the flanking genetic region among
these interfertile strains (Zuccarello and West, 1997). Furthermore, the Pacific
Mexican strains were more reproductively compatible as well as genealogically
closer to some strains from the Atlantic USA than other U.S. strains (Zuccarello
and West, 2003). They also demonstrated that the samples with the same plastid
haplotypes were sexually compatible and that those with different plastid haplo-
types were reproductively isolated, though there were some exceptions.
Although seaweeds are generally considered poor dispersers owing to short
life of spores and gametes (Destombe et al., 1990; Santelices, 1990; Shanks et al.,
2003), various seaweeds may get a lift on substrates transported long distances
(van den Hoek, 1987), such as drifting Sargassum thalli, frequently carrying many
epiphytic algae (Oliveira et al., 1979). Some red algae, such as Bostrychia,
Caloglossa, and Catenella, which are abundant in pantropical estuaries, are often
epiphytes on mangrove pneumatophores or trunks (Tanaka and Chihara, 1987),
and hence, driftwood of mangroves may facilitate distribution of these algae.
Man is undoubtedly the most potent vector of long-distance dispersal, and many
algal dispersals are presumed to have been assisted by transplanted oysters, ships’
hulls and ballast water, fishing nets, and plastic debris, or other unknown factors
(Doty, 1961; Loosanoff, 1975; Critchley and Dijkema, 1984; Carlton and Scanlon,
1985; Hay, 1990; Barnes, 2002; Flagella et al., 2007; Zuccarello et al., 2008).
However, partial incompatibility, including production of inviable progenies
and pseudocystocarps, has been mostly observed between geographically distant
populations (Table 1). Variable percentages of healthy tetrasporelings were
observed in the crossings of Digenea simplex from Caribbean coasts, mid-Atlantic
Islands, and Western Australia (Pakker et al., 1996). Despite the reduced produc-
tion of healthy tetrasporelings between the Caribbean and mid-Atlantic isolates,
they indicated close genetic distances based on RAPD analysis and similar tem-
perature tolerances, and these data suggest trans-Atlantic dispersal of this species
in the recent geological past (Pakker et al., 1996). So far, ten mating groups have
been recognized in Caloglossa leprieurii complex, and these mating groups are
fully intersterile or partially compatible (Kamiya, 2004). In this species, without
exception, reproductive isolation is completely established between the sympatri-
cally distributed mating groups, and inviable progenies or pseudocystocarps are
produced between the geographically separated groups, and never between
the sympatric groups. It is evident that allopatric speciation has a great effect on
the evolution of these algae like many terrestrial organisms, and that genetic
exchange is surely restricted between the populations in which reproductive isolation
is not fully established.
98

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
In contrast to many instances that demonstrate partial compatibility
between geographically separated populations, we have little data to suggest
partial compatibility between adjacent populations. One good example is indi-
cated by Brodie et al. (1997), who performed cross hybridizations between two
formae of Chondrus pinnulatus, f. armatus (described as a distinct species,
C. armatus, in this paper), and f. pinnulatus, whose gross morphology was
different even in the same culture condition. Forma pinnulatus was observed to
have a more northerly distribution than f. armatus, but they were found to be
sympatrically distributed in northern Japan. The results of interformae crosses
were different, from no reproductive reaction to formation of fertile F1 game-
tophytes. Their reproductive compatibilities were not correlated to their geo-
graphic distances, because f. pinnulatus female from Muroran was found to be
interfertile with f. armatus male from every locality examined, and also fertile
F1 tetrasporophytes were produced between these formae isolated from the
same locality. The two formae, however, showed a difference in the upper tem-
perature tolerance, which may probably be associated with their different dis-
tribution pattern, but the gametophytes of either formae indicated similar
reproductive phenology (Brodie et al., 1997). By considering their distinct mor-
phological differences, a prezygotic isolation mechanism such as a subtle dif-
ference in maturation timing can be established. Alternatively, gene flow
between the adjacent populations may be restricted even without any obvious
geographic barrier to dispersal. Destombe et al. (1990) showed that the mean
fertile life of spermatium in Gracilaria gracilis is about only five hours and that
spermatial dispersal is leptokurtic and limited to less than 100 m in the field.
Recent studies using molecular markers also indicate that red algal popula-
tions are highly differentiated at the level of a few to tens of kilometers (e.g.,
Intasuwan et al., 1993; Wright et al., 2000; Zuccarello et al., 2001; Zuccarello
and West, 2003; Engel et al., 2004).
5. Perspective
Genetic variation can accumulate between reproductively compatible entities
owing to external barriers or geographical separation, and the genetic distance
is mostly, but not necessarily, correlated to the reproductive affinity. Therefore,
it is important to couple the genetic data with information on reproductive com-
patibility to infer how populations differentiate, and hence, speciate. Frequent
occurrence of intraspecific reproductive isolation has been revealed by crossing
experiments with various species, and this information may give clues to eluci-
date the process of red algal speciation. Furthermore, as shown in the subsequent
paragraphs, crossing studies may be indispensable for new lines of investiga-
tions on the genetic mechanisms of red algal life history variation, phenotypic
variation, and adaptive evolution.
99

MITSUNOBU KAMIYA AND JOHN A. WEST
5.1. SPECIATION PATTERNS
Allopatric speciation is well documented in various organisms, whereas sympatric
speciation or divergence of species without geographic isolation has become
increasingly accepted as a result of theoretical works (Via, 2001). Nonetheless,
very few empirical studies for sympatric speciation have been documented
in nature (Schliewen et al., 1994; Filchak et al., 2000; Barluenga et al., 2006;
Savolainen et al., 2006). It is still difficult to prove that speciation in a given
pair of taxa has occurred in an exclusively sympatric manner, because persua-
sive cases of sympatric speciation must engage biogeographic and phylogenetic
tracks that make the existence of an allopatric phase highly likely (Jiggins,
2006). In red algae, incompletely compatible entities were usually isolated from
geographically distant populations, occasionally from adjacent populations,
and this suggests that allopatric speciation is dominant in these organisms. We
still do not have direct evidence to prove the incidence of sympatric speciation,
and remain ignorant of how frequent this process is in red algae; hence, analyses
of reproductive compatibility should be continued to unravel the pattern of red
algal speciation.
5.2. APOMIXIS
Apomixis, or reproduction without fertilization and meiosis, has been observed in
various red algal taxa, and spore recycling has been found to be relatively common
in red algae, reported from nearly 40 genera, though parthenogenesis of gametes
has been found in only a few genera (reviewed by Hawkes, 1990; West et al., 2001).
In ferns and flowering plants, apomixis and polyploidization are the outcomes of
the temporal deregulation of normal sexual reproductive pathways, sometimes
caused by interspecific hybridization (Praekelt and Scott, 2001; Park and Kato,
2003; Schranz et al., 2005). Although red algal apomixis has been recognized for a
long time, and that apomictic entities are much more dominant than sexual ones in
some species and localities (Maggs, 1988), the causes of apomixes have remained
unknown. Quite recently, apomictic tetrasporophytes of Caloglossa monosticha
were unexpectedly obtained through outcrossings between a male strain from
Australia and several female strains from Indonesia (Kamiya and West, 2008).
As no reproductive reaction or formation of pseudocystocarps were observed in
the reciprocal crosses, reproductive isolation seems to have progressed substan-
tially between the strains. As many apomictic strains of other Caloglossa species
have been isolated worldwide (West et al., 1994, 2001) and some of them were
highly heterozygous in the nuclear actin gene (M. Kamiya and J. A. West, unpub-
lished data, 2010), we presume that some parts of apomictic species originated
from such outcrossings between genetically different entities. Further crossing
experiments are required to investigate the origin of these apomicts and clarify
the genetic mechanism of apomixis.
100

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
5.3. HETEROSIS
Heterosis or hybrid vigor is a phenomenon in which an F1 hybrid has superior
performance over its parents. It has been observed in many plant species (Birchler
et al., 2003), and the utilization of heterosis is responsible for the commercial suc-
cess of plant breeding in several crops and horticultural species (Duvick, 1999).
However, there have been only a few instances suggesting macroalgal heterosis
(Hara and Akiyama, 1985; Patwary and van der Meer, 1994). For example, the
hybrid tetrasporophytes of Gelidium vagum consistently exhibited growth supe-
riority over the inbred tetrasporophytes in all growth experiments, although the
number of tetrasporangial stichidia in the hybrids were fewer than in the related
inbred lines (Patwary and van der Meer, 1994). Though the discovery of heterotic
hybrids and demonstration of heterosis usually requires considerable works, it is
important to investigate this phenomenon, because heterosis can be exploited for
developing economic seaweeds more appropriate to mariculture (see Guillemin
et al., 2008). In addition, heterosis may possibly be associated with diploid dom-
inance among populations, which have been reported in various red algae (see
Fierst et al., 2005). Diploid dominance may be attributed to the enhanced fitness
of the diploid phase, probably as a result of heterosis and/or the masking of dele-
terious recessive alleles (Guillemin et al., 2008). Heterosis is found to disappear in
the next haploid gametophyte, but can be maintained through apomixis (Bilinski
et al., 1989).
5.4 GENETIC BASIS OF ADAPTATION
Outcrossing experiments are sometimes indispensable for genetic analyses of
morphological variation and/or physiological adaptation. As most of the phe-
notypic traits of interest vary in degree and can be attributed to the interactions
between many genes, such polygenic traits do not follow patterns of Mendelian
inheritance. Elucidating the entire complement of genes related to a polygenic trait
provides the basis of understanding the effect of the genotype of an individual in
nature. Chromosomal location of quantitative trait loci (QTLs) can be inferred by
analyzing the band patterns of anonymous molecular markers, such as AFLP and
microsatellites, on the recombinant inbred lines derived from the hybrids between
the parents that show a different phenotypic trait of interest, and finding associa-
tion between markers and quantitative trait. Such a QTL mapping analysis has
been used for studying the genetic basis of adaptation and can provide clues about
the evolutionary history of populations, causes of the population differentiation,
and genetic basis of heterosis, which is still uncertain even in plants and animals
(Zeng, 2005; Garcia et al., 2008). Although we do not know any QTL mapping
studies on macroalgae, some of the outcrossing data introduced in this chapter
should be applicable to this analysis. For example, Gabrielsen et al. (2002) found in
Ceramium tenuicorne that the frequencies of some RAPD bands were correlated
101

MITSUNOBU KAMIYA AND JOHN A. WEST
to the salinity regime in their habitats. If these randomly derived markers are
closely linked to the genes that are responsible for the adaptation to different lev-
els of salinity, they may be good candidates for studying the molecular basis of
salinity tolerance.
6. Acknowledgments
We are grateful to Dr. Giuseppe C. Zuccarello for many useful comments and
critical reading of the manuscript. Various parts of the research leading to this
publication have been partially supported by grants to MK from Ministry of
Education, Science, Culture and Sport, Japan, and to JAW from the Australian
Research Council, Australian Biological Resources Study and Hermon Slade
Foundation.
7. References
Abe, T. and Masuda, M. (1998) Laurencia japonensis sp. nov. (Ceramiales, Rhodophyta). Eur. J. Phy-
col. 33: 17–24.
Aponte, N.E. and Ballantine, D.L. (1990) The life history in culture of Callithamnion boergesenii sp.
nov. (Ceramiaceae, Rhodophyta) from the Caribbean. Phycologia 29: 191–199.
Bäck, J., Collins, C. and Russell, G. (1992) Effects of salinity on growth of Baltic and Atlantic Fucus
vesiculosus. Br. Phycol. J. 27: 39–47.
Barluenga, M., Stölting, K.N., Salzburger, W., Muschick, M. and Meyer, A. (2006) Sympatric specia-
tion in Nicaraguan crater lake cichlid fish. Nature 439: 719–723.
Barnes, D.K.A. (2002) Invasions by marine life on plastic debris. Nature 416: 808–809.
Bergström, S. and Carlsson, B. (1994) River runoff to the Baltic Sea: 1950–1990. Ambio 23: 280–287.
Bergström, L. and Kautsky, L. (2006) Local adaptation of Ceramium tenuicorne (Ceramiales, Rhodo-
phyta) within the Baltic Sea. J. Phycol. 42: 36–42.
Bergström, L., Bruno, E., Eklund, B. and Kautsky, L. (2003) Reproductive strategies of Ceramium
tenuicorne near its inner limit in the brackish Baltic Sea. Bot. Mar. 46: 125–131.
Bilinski, C.A., Marmiroli, N. and Miller, J.J. (1989) Apomixis in Saccharomyces cerevisiae and other
eukayrotic micro-organisms. Adv. Microbiol. Physiol. 30: 23–52.
Birchler, J.A., Auger, D.L. and Riddle, N.C. (2003) In search of the molecular basis of heterosis. Plant
Cell 15: 2236–2239.
Bird, C.J. and McLachlan, J. (1982) Some underutilized taxonomic criteria in Gracilaria (Rhodophyta,
Gigartinales). Bot. Mar. 25: 557–562.
Bird, C.J., van der Meer, J.P. and McLachlan, J. (1982) A comment on Gracilaria verrucosa (Huds.)
Papenf. (Rhodophyta: Gigartinales). J. Mar. Biol. Ass. U.K. 62: 453–459.
Bolwell, G.P., Callow, J.A. and Evans, L.V. (1980) Fertilization in brown algae. III. Preliminary char-
acterization of putative gamete receptors from eggs and sperm of Fucus serratus. J. Cell Sci. 43:
209–224.
Boney, A.D. (1960) Observations on the spore output of some common red algae. Br. Phycol. Bull.
2: 36–37.
Boo, S.M. and Lee, I.K. (1983) A life history and hybridization of Antithamnion sparsum Tokida (Rho-
dophyta, Ceramiaceae) in culture. Kor. J. Bot. 26: 141–150.
Brawley, S.H. and Johnson, L.E. (1992) Gametogenesis, gametes and zygotes: an ecological perspec-
tive on sexual reproduction in the algae. Br. Phycol. J. 27: 233–252.
102

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
Brodie, J. and Zuccarello, G.C. (2006) Systematics of the species-rich algae: red algal classification,
phylogeny and speciation, In: T.R. Hodkinson and J.A.N. Parnell (eds.) Reconstructing the Tree
of Life: Taxonomy and Systematics of Species Rich Taxa. Systematics Association Series, CRC
Press, Boca Raton, Florida, USA, pp. 317–330.
Brodie, J., Guiry, M.D. and Masuda, M. (1993) Life history, morphology and crossability of Chondrus
ocellatus forma ocellatus and C. ocellatus forma crispoides (Gigartinales, Rhodophyta) from the
north-western Pacific. Eur. J. Phycol. 28: 183–196.
Brodie, J., Masuda, M., Mine, I. and Guiry, M.D. (1997) Two morphologically similar biological spe-
cies: Chondrus pinnulatus and C. armatus (Gigartinaceae, Rhodophyta). J. Phycol. 33: 682–698.
Carlton, J.T. and Scanlon, J.A. (1985) Progression and dispersal of an introduced alga: Codium fragile
ssp. tomentosoides (Chlorophyta) on the Atlantic coast of North America. Bot. Mar. 28: 155–
165.
Chen, C.C.-M. and Taylor, A.R.A. (1980) Investigations of distinct strains of Chondrus crispus Stackh.
II. Culture studies. Bot. Mar. 23: 441–448.
Chopin T., Bird, C.J., Murphy, C.A., Osborne, J.A., Patwary, M.U. and Floc’h, J.-Y. 1996. A molecular
investigation of polymorphism in the North Atlantic red alga Chondrus crispus (Gigartinales).
Phycol. Res. 44: 69–80.
Coyer, J.A., Peters, A.F., Stam, W.T. and Olsen, J.L. (2003) Post-ice age recolonization and differentia-
tion of Fucus serratus L. (Phaeophyceae; Fucaceae) populations in Northern Europe. Mol. Ecol.
12: 1817–1829.
Critchley, A.T. and Dijkema, R. (1984) On the presence of the introduced brown alga Sargassum
muticum, attached to commercially imported Ostrea edulis in the S.W. Netherlands. Bot. Mar.
27: 211–216.
Destombe, C., Godin, J. and Remy, J.-M. (1990) Viability and dissemination of spermatia of Gracilaria
verrucosa (Gracilariales, Rhodophyta). Hydrobiologia 204/205: 219–223.
De Wreede, R.E. and Klinger, T. (1988) Reproductive strategies in algae, In: J. Lovett-Doust and
L. Doust (eds.) Plant Reproductive Ecology: Patterns and Strategies. Oxford University Press,
Oxford, New York, USA, pp. 267–284.
Donoghue, M.J. (1985) A critique of the biological species concept and recommendations for a phylo-
genetic alternative. The Bryologist 88: 172–181.
Doty, M. (1961) Acanthophora, a possible invader of the marine flora of Hawaii. Pacific Sci. 15:
547–552.
Duvick, D.N. (1999) Heterosis: feeding people and protecting natural resources, In: J.G. Coors and S.
Pandey (eds.) Genetics and Exploitation of Heterosis in Crops. American Society of Agronomy/
Crop Science Society of America, Madison, WI, USA, pp. 19–29.
Engel, C.R. and Destombe, C. (2002) Reproductive ecology of an intertidal red seaweed, Gracilaria
gracilis: influence of high and low tides on fertilization success. J. Mar. Biol. Ass. U.K. 82:
189–192.
Engel, C.R., Destombe, C. and Valero, M. (2004) Mating system and gene flow in the red seaweed
Gracilaria gracilis: effect of haploid-diploid life history and intertidal rocky shore landscape on
fine-scale genetic structure. Heredity 92: 289–298.
Engel, C.R., Valeo, M., Lagadeuc, Y. and Destombe, C. (2002) Non-random mating in controlled
multiple-donor crosses in Gracilaria gracilis (Gracilariaceae, Rhodophyta). Eur. J. Phycol. 37:
179–190.
Engel, C.R., Wattier, R., Destombe, C. and Valero, M. (1999) Performance of non-motile male gam-
etes in the sea: analysis of paternity and fertilization success in a natural population of a red
seaweed, Gracilaria gracilis. Proc. R. Soc. Lond. Biol. Ser. B 266: 1879–1886.
Fierst, J., terHorst, C., Kübler, J.E. and Dudgeon, S. (2005) Fertilization success can drive patterns of
phase dominance in complex life histories. J. Phycol. 41: 238–249.
Filchak, K.E., Roethele, J.B. and Feder, J.L. (2000) Natural selection and sympatric divergence in the
apple maggot Rhagoletis pomonella. Nature 407: 739–742.
Flagella, M.M., Verlaque, M., Soria, A. and Buia, M.C. (2007) Macroalgal survival in ballast water
tanks. Mar. Poll. Bull. 54: 1395–1401.
103

MITSUNOBU KAMIYA AND JOHN A. WEST
Fredriksen, S., Guiry, M.D. and Rueness, J. (1994) Morphological and biosystematic studies of
Gelidium pusillum and G. pulchellum (Gelidiaceae, Rhodophyta) from Europe. Phycologia 33:
462–470.
Furnari, G., L’Hardy-Halos, M.-T., Rueness, J. and Serio, D. (1998) On the conspecificity of
(Aglaothamnion tenuissimum and A. byssoides (ceramiaceae, Rhodophyta). Taxon 47: 843–849.
Gabrielsen, T.M., Brochmann, C. and Rueness, J. (2002) The Baltic Sea as a model system for study-
ing postglacial colonization and ecological differentiation, exemplified by the red alga Ceramium
tenuicorne. Mol. Ecol. 11: 2083–2095.
Gabrielsen, T.M., Brochmann, C. and Rueness, J. (2003) Phylogeny and interfertility of North Atlan-
tic populations of ‘Ceramium strictum’ (Ceramiales, Rhodophyta): how many species? Eur. J.
Phycol. 38: 1–13.
Garbary, D. (1988) Interoceanic hybridization in fully corticated Ceramium isolates (Rhodophyta)
from Nova Scotia and Washington. Kor. J. Phycol. 3: 89–93.
Garcia, A.A.F., Wang, S., Melchinger, A.E. and Zeng, Z.-B. (2008) Quantitative trait loci mapping and
the genetic basis of heterosis in maize and rice. Genetics 180: 1707–1724.
Goff, L.J. and Coleman, A.W. (1984) Elucidation of fertilization and development in a red alga by
quantitative DNA microspectrofluorometry. Devel. Biol 102: 173–194.
Goff, L.J. and Coleman, A.W. (1990) DNA: microspectrofluorometric studies, In: K.M. Cole and R.G.
Sheath (eds.) Biology of the Red Algae. Cambridge University Press, New York, USA, pp. 43–71.
Goodenough, U.W. and Adair, W.S. (1989) Recognition proteins of Chlamydomonas reinhardtii (Chlo-
rophyceae), In: A.W. Coleman, L.J. Goff and J.R. Stein-Taylar (eds.) Algae as Experimental Sys-
tems. Alan R. Liss, Inc., New York, USA, pp. 171–185.
Gordon, R. and Brawley, S.H. (2004) Effects of water motion on propagule release from algae with
complex life histories. Mar. Biol. 145: 21–29.
Graham, L.E. and Wilcox, L.W. (2000) Algae. Prentice Hall, New Jersey, USA, pp. 640.
Guillemin, M.-L., Faugeron, S., Destombe, C., Viard, F., Correa, J.A. and Valero, M. (2008) Genetic
variation in wild and cultivated populations of the haploid-diploid red alga Gracilaria chilensis:
how farming practices favor asexual reproduction and heterozygosity. Evolution 62: 1500–1519.
Guiry, M.D. (1981) Chondrus crispus Stackhouse “T4” is a male clone. Phycologia 20: 438–439.
Guiry, M.D. (1984) Structure, life history and hybridisation of Atlantic Gigartina teedii (Rhodophyta)
in culture. Br. Phycol. J. 19: 37–55.
Guiry, M.D. (1992) Species concepts in marine red algae, In: F.E. Round and D.J. Chapman (eds.)
Progress in Phycological Research, Vol. 8. Biopress, Ltd., Bristol, UK, pp. 251–278.
Guiry, M.D. and Freamhainn, M.T. (1986) Biosystematics of Gracilaria foliifera (Gigartinales, Rho-
dophyta). Nord. J. Bot. 5: 629–637.
Guiry, M.D. and West, J.A. (1983) Life history and hybridization studies on Gigartina stellata and
Petrocelis cruenta (Rhodophyta) in the North Atlantic. J. Phycol. 19: 474–494.
Guiry, M.D., Tripodi, G. and Lüning, K. (1987) Biosystematics, genetics and upper temperature tol-
erance of Gigartina teedii (Rhodophyta) from the Atlantic and Mediterranean. Helgol. Meere-
sunters. 41: 283–295.
Hara, M. and Akiyama, K. (1985) Heterosis in growth of Undaria pinnatifida (Harvey) Suringar. Bull.
Tohoku Reg. Fish. Res. Lab. 47: 47–50 (in Japanese with English abstract).
Hawkes, M.W. (1990) Reproductive strategies, In: K.M. Cole and R.G. Sheath (eds.) Biology of the
Red Algae. Cambridge University Press, New York, USA, pp. 455–476.
Hay, C.H. (1990) The dispersal of sporophytes of Undaria pinnatifida by coastal shipping in New Zea-
land, and implications for further dispersal of Undaria in France. Br. Phycol. J. 25: 301–313.
Hoffmann, A.J. (1987) The arrival of seaweed propagules at the shore: a review. Bot. Mar. 30: 151–165.
Hommersand, M.H. and Fredericq, S. (1990) Sexual reproduction and cystocarp development, In:
K.M. Cole and R.G. Sheath (eds.) Biology of the Red Algae. Cambridge University Press, New
York, USA, pp. 43–71.
Hu, Z., Zeng, X., Critchley, A.T., Morrell, S.L. and Duan, D. (2007) Phylogeography of the Northern
Atlantic species Chondrus crispus (Gigartinales, Rhodophyta) inferred from nuclear rDNA inter-
nal transcribed spacer sequences. Hydrobiologia 575: 315–327.
104

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
Intasuwan, S., Gordon, M.E., Daugherty, C.H. and Lindsay, G.C. (1993) Assessment of allozyme
variation among New Zealand populations of Gracilaria chilensis (Gracialiares, Rhodophyta)
using starch-gel electrophoresis. Hydrobiologia 260/261: 159–165.
Jiggins, C.D. (2006) Sympatric Speciation: Why the controversy? Curr. Biol. 16: R333–R334.
Johannesson, K. and André, C. (2006) Life on the margin – genetic isolation and diversity loss in a
peripheral marine ecosystem, the Baltic Sea. Mol. Ecol. 15: 2013–2029.
Johansson, G., Sosa, P.A. and Snoeijs, P. (2003) Genetic variability and level of differentiation in
North Sea and Baltic Sea populations of the green alga Cladophora rupestris. Mar. Biol. 142:
1019–1027.
Jones, J.L., Callow, J.A. and Green, J.R. (1988) Monoclonal antibodies to sperm surface antigens of
the brown alga Fucus serratus exhibit region- , gamete- , species- , and genus-preferential binding.
Planta 176: 298–306.
Jones, J.L., Callow, J.A. and Green, J.R. (1990) The molecular nature of Fucus serratus sperm surface
antigens recognised by monoclonal antibodies FS1 to FS12. Planta 182: 64–71.
Kamiya, M. (2004) Speciation and biogeography of the Caloglossa leprieurii complex (Delesseriaceae,
Rhodophyta). J. Plant Res. 117: 421–428.
Kamiya, M. and Kawai, H. (2002) Dependence of the carposporophyte on the maternal gametophyte
in three Ceramiacean algae (Rhodophyta), with respect to carposporophyte development, spore
production and germination success. Phycologia 41: 107–115.
Kamiya, M. and West, J.A. (2008) Origin of apomictic red algae: outcrossing studies of different
strains in Caloglossa monosticha (Ceramiales, Rhodophyta). J. Phycol. 44: 977–984.
Kamiya, M., Tanaka, J. and Hara, Y. (1995) A morphological study and hybridization analysis of
Caloglossa leprieurii (Ceramiales, Rhodophyta) from Japan, Singapore and Australia. Phycol.
Res. 43: 81–91.
Kamiya, M., Tanaka, J. and Hara, Y. (1997) Comparative morphology, crossability, and taxonomy
within the Caloglossa continua (Delesseriaceae, Rhodophyta) complex from the western Pacific.
J. Phycol. 33: 97–105.
Kamiya, M., Tanaka, J., King, R.J., West, J.A., Zuccarello, G.C. and Kawai, H. (1999) Reproductive
and genetic distinction between broad and narrow entities of Caloglossa continua (Delesseriaceae,
Rhodophyta). Phycologia 38: 356–367.
Kamiya, M., West, J.A., King, R.J., Zuccarello, G.C., Tanaka, J. and Hara, Y. (1998) Evolutionary
divergence in the red algae Caloglossa leprieurii and C. apomeiotica. J. Phycol. 34: 361–370.
Kamiya, M., West, J.A., Zuccarello, G.C. and Kawai, H. (2000) Caloglossa intermedia sp. nov. (Rho-
dophyta) from the western Atlantic coast: molecular and morphological analyses with special
reference to C. leprieurii and C. monosticha. J. Phycol. 36: 411–420.
Kamiya, M., Zuccarello, G.C. and West, J.A. (2003) Evolutionary relationships of the genus caloglossa
(Delesseriaceae, Rhodophyta) inferred from large-subunit ribosomal RNA gene sequences, mor-
phological evidence and reproductive compatiblity, with description of a new species from
Guatemala, Phycologia 42: 478–497.
Kapraun, D.F. (1977) Asexual propagules in the life history of Polysiphonia ferulacea (Rhodophyta,
Ceramiales). Phycologia 16: 417–426.
Kim, G.H. and Fritz, L. (1993) Gamete recognition during fertilization in a red alga, Antithamnion
nipponicum. Protoplasma 174: 69–73.
Kim, S.-H. and Kim, G.H. (1999) Cell-cell recognition during fertilization in the red algae, Aglaotham-
nion oosumiense (Ceramiaceae, Rhodophyta). Hydrobiologia 398/399: 81–89.
Kim, G.H., Lee, I.K. and Fritz, L. (1996) Cell–cell recognition during fertilization in a red alga, Anti-
thamnion sparsum (Ceramiaceae, Rhodophyta). Plant Cell Physiol. 37: 621–628.
Kim, G.H., Shin, J.B., Klochkova, T.A., West, J.A. and Zuccarello, G.C. (2008) The utility of pro-
teomics in algal taxonomy: Bostrychia radicans/B. moritziana (Rhodomelaceae, Rhodophyta) as
a model study. J. Phycol. 44: 1519–1528.
Kristiansen, A.A., Pedersen, P.M. and Moseholm, L. (1994) Salinity-temperature effects on growth
and reproduction of Scytosiphon lomentaria (Fucophyceae) along the salinity gradient in Danish
waters. Phycologia 33: 444–454.
105

MITSUNOBU KAMIYA AND JOHN A. WEST
Kudo, T. and Masuda, M. (1986) A taxonomic study of Polysiphonia japonica Harvey and P. akke-
shiensis Segi (Rhodophyta). Jap. J. Phycol. 34: 293–310.
Kugrens, P. and West, J.A. (1972) Synaptonemal complexes in red algae. J. Phycol. 8: 187–191.
Lee, J.H., Waffenschmidt, S., Small, L. and Goodenough, U. (2007) Between-species analysis of short-
repeat modules in cell wall and sex-related hydroxyproline-rich glycoproteins of Chlamydomonas.
Plant Physiol. 144: 1813–1826.
L’Hardy-Halos, M.-Th. and Rueness, J. (1990) Comparative morphology and crossability of related
species of Aglaothamnion (Rhodophyta). Phycologia 29: 351–366.
Loosanoff, V.L. (1975) Introduction of Codium in New England waters. Fish. Bull. 73: 215–218.
Ma, J.H. and Miura, A. (1984) Observations of the nuclear division in the conchospores and their
germlings in Porphyra yezoensis Ueda. Jap. J. Phycol. 32: 373–378 (in Japanese with English
abstract).
Maggs, C.A. (1988) Intraspecific life history variability in the Florideophycidae (Rhodophyta). Bot.
Mar. 31: 465–490.
Magruder, W.H. (1984) Specialized appendages on spermatia from the red alga Aglaothamnion neglec-
tum (Ceramiales, Ceramiaceae) specifically bind with trichogynes. J. Phycol. 20: 436–440.
Masuda, M. and Kogame, K. (1998) Crossability between Hawaiian and Japanese populations of
Ahnfeltiopsis concinna (Gigartinales, Rhodophyta). Bot. Mar. 41: 243–247.
Masuda, M., West, J.A. and Kurogi, M. (1987) Life history studies in culture of a Mastocarpus species
(Rhodophyta) from central Japan. J. Fac. Sci., Hokkaido Univ. Ser. V (Botany) 14: 11–38.
Masuda, M., West, J.A., Ohno, Y. and Kurogi, M. (1984) Comparative reproduction patterns in cul-
ture of different Gigartina subgenus Mastocarpus and Petrocelis populations from Northern
Japan. Bot. Mag. Tokyo 97: 107–125.
Mathieson, A.C., Norton, T.A. and Neushul, M. (1981) The taxonomic implications of genetic and
environmentally induced variations in seaweed morphology. Bot. Rev. 47: 313–347.
Mayr, E. (1942) Systematics and the Origin of Species from the Viewpoint of a Zoologist. Columbia
University Press, New York, pp. 334.
Mayr, E. (1954) Geographic speciation in tropical echinoids. Evolution 8: 1–18.
McLachlan, J., van der Meer, J.P. and Bird, N.L. (1977) Chromosome numbers of Gracilaria foli-
ifera and Gracilaria sp. (Rhodophyta) and attempted hybridizations. J. Mar. Biol. Ass. U.K. 57:
1137–1141.
Middelboe, A.L., Sand-Jensen, K. and Brodersen, K. (1997) Patterns of macroalgal distribution in the
Kattegat-Baltic region. Phycologia 36: 208–219.
Mine, I. and Tatewaki, M. (1994) Attachment and fusion of gametes during fertilization of Palmaria
sp. (Rhodophyta). J. Phycol. 30: 55–66.
Miura, A, Fu, P.F. and Shin, J-A. (1992) Interspecific cross experiments between Porphyra yezoensis
Ueda and P. tenera Kjellman (Bangiales, Rhodophyta) by using pigmentation variants. J. Tokyo
Univ. Fish. 79: 103–120 (in Japanese with English abstract).
Molenaar, F.J. and Breeman, A.M. (1994) Ecotypic variation in Phyllophora pseudoceranoides (Rho-
dophyta) ensures winter reproduction throughout its geographic range. J. Phycol. 30: 392–402.
Molenaar, F.J., Breeman, A.M. and Venekamp, L.A.M. (1996) Ecotypic variation in Cystoclonium
purpureum (Rhodophyta) synchronizes life history events in different regions. J. Phycol. 32:
516–525.
Nečas, J. (1981) Dependence of the gametogenesis induction, zygote formation and their germina-
tion on the culture density of the homothallic alga Chlamydomonas geitleri Ettl. Biol. Plant. 23:
278–284.
Norton, T.A. (1992) Dispersal by macroalgae. Br. Phycol. J. 27: 293–301.
Ohme, M. and Miura, A. (1988) Tetrad analysis in conchospore germlings of Porphyra yezoensis
(Rhodophyta, Bangiales). Plant Sci. 57: 135–140.
Oliveira, E.C., Ugadim, Y. and De Paula, E.J. (1979) Associated epibiota on Sargassum floating on the
waters of the Brazilian current – biogeographical remarks. Bolm. botanica, Univ. São Paulo 7:
5–9 (in Portuguese with English abstract).
Olsen, J.L., Stam, W.T., Coyer, J.A. et al. (2004) North Atlantic phylogeography and large-scale popu-
lation differentiation of the seagrass Zostera marina L. Mol. Ecol. 13: 1923–1941.
106

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
Pakker, H., Klerk, H., van Campen, J.H., Olsen, J.L. and Breeman, A.M. (1996) Evolutionary and
ecological differentiation in the pantropical to warm-temperate seaweed Digenea simplex (Rho-
dophyta). J. Phycol. 32: 250–257.
Palumbi, S.R. (1994) Genetic divergence, reproductive isolation and marine speciation. Annu. Rev.
Ecol. Syst. 25: 547–572.
Park, C.-H. and Kato, M. (2003) Apomixis in the interspecific triploid hybrid fern Cornopteris chris-
tenseniana (Woodsiaceae). J. Plant Res. 116: 93–103.
Patwary, M.U. and van der Meer, J.P. (1994) Application of RAPD markers in an examination of
heterosis in Gelidium vagum (Rhodophyta). J. Phycol. 30: 91–97.
Pickett-Heaps, J.D. and West, J.A. (1998) Time-lapse video observations on sexual plasmogamy in the
red alga Bostrychia. Eur. J. Phycol. 33: 43–56.
Plastino, E.M. and de Oliveira, E.C. (1988) Sterility barriers among species of Gracilaria (Rhodo-
phyta, Gigartinales) from the São Paulo littoral, Brazil. Br. Phycol. J. 23: 267–271.
Polanshek, A.R. and West, J.A. (1975) Culture and hybridization studies on Petrocelis (Rhodophyta)
from Alaska and California. J. Phycol. 11: 434–439.
Polanshek, A.R. and West, J.A. (1977) Culture and hybridization studies on Gigartina papillata (Rho-
dophyta). J. Phycol. 13: 141–9.
Praekelt, U. and Scott, R. (2001) Induction of apomixis in sexual plants by mutagenesis, In: Y. Savidan,
J. G. Carman and T. Dresselhaus (eds.) The Flowering of Apomixis: from Mechanisms to Genetic
Engineering. Mexico, D.F.: CIMMYT, IRD, European Commission DG VI, pp. 212–228.
Reusch, T.B.H. (2002) Microsatellites reveal high population connectivity in eelgrass (Zostera marina)
in two contrasting coastal areas. Limnol.Oceanogr. 47: 78–85.
Reusch, T.B.H., Stam, W.T. and Olsen, J.L. (2000) A microsatellite-based estimation of clonal diversity
and population subdivision in Zostera marina, a marine flowering plant. Mol. Ecol. 9: 127–140.
Rice, E.L. and Bird, C.J. (1990) Relationships among geographically distant populations of Gracilaria
verrucosa (Gracilariales, Rhodophyta) and related species. Phycologia 29: 501–510.
Richards, J.S. and Sommerfeld, M.R. (1974) Gamete activity in mating strains of Chlamydomonas
eugametos. Arch. Mikrobiol. 98: 69–75.
Richerd, S., Destombe C., Cuguen, J. and Valero, M. (1993) Variation of reproductive success in a
haplo-diploid red alga, Gracilaria verrucosa: effects of parental identities and crossing distance.
Am. J. Bot. 80: 1379–1391.
Rietema, H. (1991) Evidence for ecotypic divergence between Phycodrys rubens populations from the
Baltic Sea and North Sea. Bot. Mar. 34: 375–381.
Rietema, H. (1993) Ecotypic differences between Baltic and North-Sea populations of Delesseria san-
guinea and Membranoptera alata. Bot. Mar. 36: 15–21.
Rietema, H. (1995) Ecoclinal variation in Rhodomela confervoides along a salinity gradient in the
North Sea and Baltic Sea. Bot. Mar. 38: 475–479.
Rueness, J. (1973) Speciation in Polysiphonia (Rhodophyceae, Ceramiales) in view of hybridization
experiments: P. hemisphaerica and P. boldii. Phycologia 12: 107–109.
Rueness, J. (1978) Hybridization in red algae. In: D. E. G. Irvine and J. H. Price (eds.) Modern Approaches
to the Taxonomy of Red and Brown Algae, Vol.10. Academic Press, London, UK, pp. 247–262.
Rueness, J. and Kornfeldt, R-A. (1992) Ecotypic differentiation in salinity responses of Ceramium
strictum (Rhodophyta) from Scandinavian waters. Sarsia 77: 207–212.
Rueness, J. and Rueness, M. (1975) Genetic control of morphogenesis in two varieties of Antithamnion
plumula (Rhodophyceae, Ceramiales). Phycologia 14: 81–85.
Russell, G. (1985) Recent evolutionary changes in the algae of the Baltic Sea. Br. Phycol. J. 20: 87–104.
Russell, G. (1994) A Baltic variant of Pilayella littoralis (Algae, Fucophyceae). Ann. Bot. Fennici 31:
127–138.
Santelices, B. (1990) Patterns of reproduction, dispersal and recruitment in seaweeds. Oceanogr. Mar.
Biol. 28: 177–276.
Santelices, B. (2002) Recent advances in fertilization ecology of macroalgae. J. Phycol. 38: 4–10.
Savolainen, V., Anstett, M.-C., Lexer, C., Hutton, I, Clarkson, J.J, Norup, M. V., Powell, M.P., Spring-
ate, D., Salamin, N. and Baker, W.J. (2006) Sympatric speciation in palms on an oceanic island.
Nature 441: 210–213.
107

MITSUNOBU KAMIYA AND JOHN A. WEST
Schliewen, U.K., Tautz, D. and Pääbo, S. (1994) Sympatric speciation suggested by monophyly of
crater lake cichlids. Nature 368: 629–632.
Schranz M.E., Dobeš, C., Koch, M.A. and Mitchell-Olds T. (2005) Sexual reproduction, hybridization,
apomixis, and polyploidization in the genus Boechera (Brassicaceae). Am. J. Bot. 92: 1797–1810.
Searles, R.B. (1980) The strategy of the red algal life history. Am. Nat. 115: 113–120.
Serrão, E.A., Kautsky, L. and Brawley, S.H. (1996) Distributional success of the marine seaweed Fucus
vesiculosus L. in the brackish Baltic Sea correlates with osmotic capabilities of Baltic gametes.
Oecologia 107: 1–12.
Shanks, A.L., Grantham, B.A. and Carr, M.H. (2003) Propagule dispersal distance and the size and
spacing of marine reserves. Ecol. Appl. 13: S159–S169. (supplement)
Shimizu, A., Morishima, K., Kobayashi, M., Kunimoto, M. and Nakayama, I. (2008) Identification of
Porphyra yezoensis (Rhodophyta) meiosis by DNA quantification using confocal laser scanning
microscopy. J. Appl. Phycol. 20: 83–88.
Spencer, K.G., Yu, M-H., West, J.A. and Glazer, A.N. (1981) Phycoerythrin and interfertility patterns
in Callithamnion (Rhodophyta) isolates. Br. Phycol. J. 16: 331–343.
Starr, R.C. and Jaenicke, L. (1974) Purification and characterization of the hormone initiating sexual
morphogenesis in Volvox carteri f. nagariensis Iyengar. Proc. Natl. Acad. Sci. USA 71: 1050–1054.
Sundene, O. (1975) Experimental studies on form variation in Antithamnion plumula (Rhodophyceae).
Norw. J. Bot. 22: 35–42.
Suto, S. (1963) Intergeneric and interspecific crossings of the lavers (Porphyra). Bull. Jap. Soc. Sci.
Fish. 29: 739–748.
Tanaka, J. and Chihara, M. (1987) Species composition and vertical distribution of macroalgae in brack-
ish waters of Japanese mangrove forests. Bull. Natl. Sci. Mus., Tokyo, Ser. B (Botany) 13: 141–150.
Tanaka, J. and Kamiya, M. (1993) Reproductive structure of Caloglossa ogasawaraensis Okamura
(Ceramiales, Rhodophyceae) in nature and culture. Jap. J. Phycol. 41: 113–121.
Thomas, D.N, Collins, J.C. and Russell, G. (1990) Interpopulational differences in salt tolerances of
two Cladophora species. Est. Coast. Shelf Sci. 30: 201–206.
Togashi, T. and Cox, P.A. (2004) Phototaxis and the evolution of isogamy and ‘slight anisogamy’ in
marine green algae: insights from laboratory observations and numerical experiments. Bot. J.
Linn. Soc. 144: 321–327.
Tsuchikane, Y., Fukumoto, R., Akatsuka, S., Fujii, T. and Sekimoto, H. (2003) Sex pheromones that
induce sexual cell division in the Closterium peracerosum-strigosum-littorale complex (Charo-
phyta). J. Phycol. 39: 303–309.
Valatka, S., Mäkinen, A. and Yli-Mattila, T. (2000) Analysis of genetic diversity of Furcellaria lumbricalis
(Gigartinales, Rhodophyta) in the Baltic Sea by RAPD-PCR technique. Phycologia 39: 109–117.
van den Hoek, C. (1987) The possible significance of long-range dispersal for the biogeography of
seaweeds. Helgol. Meersunters. 41: 261–272.
van der Meer, J.P. (1986) Genetic contributions to research on seaweeds. In: F.E. Round and D.J. Chap-
man (eds.) Progress in Phycological Research, Vol. 4. Biopress, Bristol, UK, pp.1–38.
van der Meer, J.P. (1987) Experimental hybridization of Palmaria palmata (Rhodophyta) from the
northeast and northwest Atlantic Ocean. Can. J. Bot. 65: 1451–1458.
van Oppen, M.J.H., Draisma, S.G.A., Olsen, J.L. and Stam, W.T. (1995a) Multiple trans-Arctic pas-
sages in the red alga Phycodrys rubens: evidence from nuclear rDNA ITS sequences. Mar. Biol.
123: 179–188.
van Oppen, M.J.H., Olsen, J.L. and Stam, W.T. (1995b)Genetic variation within and among North
Atlantic and Baltic populations of the benthic alga Phycodrys rubens (Rhodophyta). Eur. J.
Phycol. 30: 251–260.
Via, S. (2001) Sympatric speciation in animals: the ugly duckling grows up. Trends Ecol. Evol. 16: 381–390.
Wang, J., Dai, J.X. and Zhang, Y.T. (2006) Nuclear division of the vegetative cells, conchosporangial
cells and conchospores of Porphyra yezoensis (Bangiales, Rhodophyta). Phycol. Res. 54: 201–207.
West, J.A. and McBride, D.L. (1999) Long-term and diurnal carpospore discharge patterns in the
Ceramiaceae, Rhodomelaceae and Delesseriaceae (Rhodophyta). Hydrobiologia 398/399:
101–113.
108

INVESTIGATIONS ON REPRODUCTIVE AFFINITIES IN RED ALGAE
West, J.A., Polanshek, A.R. and Shevlin, D.E. (1978) Field and culture studies on Gigartina agardhii
(Rhodophyta). J. Phycol. 14: 416–426.
West, J.A., Zuccarello, G.C. and Kamiya, M. (2001) Reproductive patterns of Caloglossa species
(Delesseriaceae, Rhodophyta) from Australia and New Zealand: multiple origins of asexuality in
C. leprieurii. Literature review on apomixis, mixed-phase, bisexuality and sexual compatiblility.
Phycol. Res. 49: 183–200.
West, J.A., Zuccarello, G.C. and Karsten, U. (1996) Reproductive biology of Stictosiphonia hookeri
(Rhodomelaceae, Rhodophyta) from Argentina, Chile, South Africa and Australia in laboratory
culture. Hydrobiologia 326/327: 277–282.
West, J.A., Zuccarello, G.C., Pedroche, F.F. and Karsten, U. (1994) Caloglossa apomeiotica sp. nov.
(Ceramiales, Rhodophyta) from Pacific México. Bot. Mar. 37: 381–390.
Wilce, R.T. and Sears, J.R. (1991) Schmitzia sanctae-crucis, new species (Calosiphoniaceae, Rhodo-
phyta) and a novel nutritive development to aid in zygote nucleus amplification. Phycologia 30:
151–169.
Wilson, S.M., Pickett-Heaps, J.D. and West, J.A. (2002) Fertilization and the cytoskeleton in the red
alga Bostrychia moritziana (Rhodomelaceae, Rhodophyta). Eur. J. Phycol. 37: 509–522.
Wilson, S.M., West, J.A. and Pickett-Heaps, J.D. (2003) Time-lapse videomicroscopy of fertilisation
and the actin cytoskeleton in Murrayella periclados. (Rhodomelaceae, Rhodophyta). Phycologia
42: 638–645.
Wright, J.T., Zuccarello, G.C., and Steinberg, P.D. (2000) Genetic structure of the subtidal red alga
Delisea pulchra. Mar. Biol. 136: 439–448.
Yamamoto, H. and Sasaki, J. (1987) Crossing experiments between populations of so-called Gracilaria
verrucosa (Huds.) Papenfuss from two localities, Shinori and Kikonai in Hokkaido. Bull. Fac.
Fish. Hokkaido Univ. 38: 335–338.
Yamamoto, H. and Sasaki, J. (1988) Interfertility between so-called Gracilaria verrucosa (Huds.) Papen-
fuss and G. vermiculophylla (Ohmi) Papenfuss in Japan. Bull. Fac. Fish. Hokkaido Univ. 39: 1–3.
Yoon, H.Y. and West, J. (1990) Comparative morphology and hybridization of Polysiphonia acuminata
from California and P. japonica var. japonica from Korea. Kor. J. Phycol. 5: 51–56.
Zeng, Z.-B. (2005) QTL mapping and the genetic basis of adaptation: recent developments. Genetica
123: 25–37.
Zuccarello, G.C. and West, J.A. (1995) Hybridization Studies in Bostrychia. I: B. radicans (Montagne)
Montagne (Rhodomelaceae, Rhodophyta) from Pacific and Atlantic North America. Phycol.
Res. 43: 233–240.
Zuccarello, G.C. and West, J.A. (1997) Hybridization studies in Bostrychia: 2. Correlation of crossing
data and plastid DNA sequence data within B. radicans and B. moritziana (Ceramiales, Rhodo-
phyta). Phycologia 36: 293–304.
Zuccarello, G.C. and West, J.A. (2003) Multiple cryptic species: molecular diversity and reproductive
isolation in the Bostrychia radicans/B. moritziana complex (Rhodomelaceae, Rhodophyta) with
focus on North American isolates. J. Phycol. 39: 948–959.
Zuccarello, G.C., Sandercock, B. and West, J.A. (2002) Diversity within red algal species: variation
in world-wide samples of Spyridia filamentosa (Ceramiaceae) and Murrayella periclados (Rho-
domelaceae) using DNA markers and breeding studies. Eur. J. Phycol. 37: 403–417.
Zuccarello, G.C., West, J.A. and King, R.J. (1999) Evolutionary divergence in the Bostrychia moritziana/
B. radicans complex (Rhodomelaceae, Rhodophyta): molecular and hybridization data. Phyco-
logia 38: 234–144.
Zuccarello, G.C., Yeates, P.H., Wright, J.T. and Bartlett, J. (2001) Population structure and physiologi-
cal differentiation of haplotypes of Caloglossa leprieurii (Rhodophyta) in a mangrove intertidal
zone. J. Phycol. 37: 235–244.
Zuccarello, G.C., West, J.A. and Kikuchi, N. (2008) Phylogenetic relationships within the Stylone-
matales (Stylonematophyceae, Rhodophyta): biogeographic patterns do not apply to Stylonema
alsidii. J. Phycol. 44: 384–393.
109