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Earthworms are generally cross-fertilization hermaphrodites, but up to 40% of the species can be parthenogenetic. In simultaneous hermaphrodites, a trade-off between male and female sexual functions is expected because the two sexes share limited resources from the same individual. In this chapter, several issues regarding sexual selection such as the role of spermathecae, copulatory behavior, allohormone injection, or adjustment of the donated sperm volume are reviewed. Parthenogenesis is present in some families as Lumbricidae, but is lacking in others. Parthenogenetic reproduction in earthworms is generally automictic and thelytokous, although apomixis and pseudogamy have been occasionally described. This kind of reproduction is poorly understood due to some background limitations such as the species concept in parthenogens or its possible origin, which are discussed in this chapter.
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Chapter 5
Reproduction of Earthworms: Sexual Selection
and Parthenogenesis
az Cosı
n, Marta Novo, and Rosa Ferna
5.1 Introduction
Earthworms are generally considered to be cross-fertilization hermaphrodites (i.e.,
using reciprocal insemination, transferring, and receiving sperm in the same copu-
lation). Although not all earthworms use this reproductive strategy, the best known
species, Lumbricus terrestris, is a cross-fertilization hermaphrodite and this strat-
egy seems to be the most widespread in earthworms. Nevertheless, cases of self-
fertilization have been reported in earthworms; Domı
nguez et al. (2003) discussed
that Eisenia andrei individuals bend themselves, allowing their spermathecal pores
to contact the ventral zone of their clitellum. The sperm is then transported from the
male pores to the spermathecae. This finding explains why 33% of isolated indivi-
duals in this study produced viable cocoons.
However, hermaphroditism is not the only reproductive mechanism and
more parthenogenetic earthworms are being discovered all the time, most of
which a re polyploid. Parthenogenetic reproduction is very frequent in the family
Lumbricidae, with more than 30 parthenogenetic species occurring in North
America (Reynolds 1974). Parthenogenesis has also been reported in families
such as Megascolecids, but has not been observed in other families, including
“Asexual” reproduction by means of bipartition, stolonisation, budding, or
similar processes has not been observed in earthworms and their ability to regener-
ate is limited. There are several reproductive models: discontinuous, sem icontin-
uous, or continuous. In Hormogaster elisae, male and female gametogenesis are
synchronized, beginning in autumn and ending in the summer. Male funnels are full
of spermatozoa and the spermathecae contain spermatozoa throughout the year , but
az Cosı
n(*), M. Novo, and R. Ferna
Departamento de Zoologı
a, Facultad de Biologı
a, Universidad Complutense de Madrid, Ciudad
Universitaria, 28040 Madrid, Spain
A. Karaca (ed.), Biology of Earthworms , Soil Biology 24,
DOI 10.1007/978-3-642-14636-7_5,
Springer-Verlag Berlin Heidelberg 2011
two peaks of reproduction have been observed, with the largest peak occurring in
the spring and the second peak occurring in autumn (Garvı
n et al. 2003).
An excellent description of the earthworm reproductive system can be found in
general zoology volumes and monographs such as Jamieson (2006), so it will be
only succinctly described in the present chapter. Earthworms are usually hermaph-
rodites in which the testes and ovaries are accompanied by a series of organs with a
male or female function. The female components typically include the ovaries
(generally one pair in the 13th segment), ovisacs (in the 14th segment), oviducts,
female pores (in the 14th segment), and spermathecae (of variable position and
number). Male components typically include the testes and male funnels (in most
cases, there are two pairs in the 10th and 11th segments and singularly a single pair
in the 11th segment), seminal vesicles (of variable number, with 2–4 occurring in
segments 9–12), deferent ducts, and male pores surrounde d by atrial glands that are
more or less developed. Other organs, such as testicular sacs (Lumbricus and
Octolasium), accessory glands (prostates), or the thecal glands associated with the
spermathecae, may also be present.
Some of the external reproductive organs, such as the clitellum, tubercula
pubertatis, and sexual papillae, are developed at sexua l maturity. The sexual
papillae include modified genital chaetae and chaetal glands, which could be used
to inject substances into the partner (see Sect. 8.2.2).
The union during copulation, which could last between 69 and 200 min in
L. terrestris, is secured by tubercula and quetae. Copulation can occur at the surface
in epigeic and anecic earthworms, which increases the depredation risk, and also
occurs in deeper layers of the soil in the case of endogeic species. The more
primitive type of copulation seems to be a simple juxtaposition of the male pores
of one individual and the spermathecal pores of the other, with the dir ect transfer of
spermatozoa. The presence of a penis has been observed in some cases, which in
reality seems to be just an elevated papilla, as in the case of some Pheretima
In most of the species in the Lumbricidae family and in other families, the
clitellum moves backwards and seminal groves are developed from the male pores
to the tubercula pubertatis. Spermatozoa flow through the seminal grove s to get into
the partner’s spermathecae pores. Details of sperm transfer are not well known with
the exception of a few species such as Pheretima sp., in which, according to Tembe
and Dubash (1961), the sperm appears to be transferred sequentially, passing first to
the anterior spermathecae and later to the posterior ones.
(1975) indicated that spermatophores have been observed in more than
20 species of lumbricids. Spermatophores are small capsules that adhere to the body
wall and can be iridescent and full of spermatozoa. Their function is not clear. It has
been suggested that the spermatophores may play a role in sperm transfer (Edwards
and Bohlen 1996), thus avoiding sperm digestion in the spermathecae and fertiliz-
ing the ova during cocoon formation Michiels (1998). Nevertheless, Monroy et al.
(2003) showed that spermatophores have no effect on the reproductive success of
Eisenia fetida and were not able to demonstrate the specific function of these
70 D.J.
az Cosı
n et al.
Complex prec opulatory behaviors have been describ ed in partner selection in
some species, including L. terrestris, in which individuals perform visits to their
neighbors’ burrows (Nuutinen and Butt 1997; Michiels et al. 2001, see Sect. 8.2.1).
Development is direct in earthworms. Fertilization occurs within cocoon s and one
or more juveniles are produced for each cocoon.
The presence of parthenogenesis in earthworms was first observed many years
ago, thanks to the contributions of authors such as Omodeo (1951), Casel lato
(1987), Jaenicke and Selander (1979) and Victorov (1997), among others.
Reynolds (1974) pointed out that in North America 35 species are anphimictic,
11 probably sexual, 4 facultative parthenogenetic, 1 possibly parthenogenetic, and
30 parthenogenetic. Casellato (1987 ) cited 25 parthenogenetic species or subspe-
cies (12 of which had even ploidy numbers and 13 of which showed odd ploidy) and
Victorov (1997) pointed out that in Russia, the number of polyploids almost equals
the number of diploids, with a ratio of 46 polyploids: 52 diploids. He observed that
polyploids (in cases of sympatry) tend to occupy the margins of the distribution
areas. According to Edwards and Bohlen (1996), the association between partheno-
genesis and high polyploidy in earthworms produces an unexpected level of
heterozygosity, an advantageous condition that provides resistance to environmen-
tal stress.
5.2 Sexual Selection in Cross-Fertilization Earthworms
In simultaneous hermaphrodites, a trade-off between male and female sexual
functions is expected becau se the two sexes share limited reso urces from the
same individual. In addition, the strategy that maximizes fitness is different for
the male and female functions. This has been explained previously by Bateman
(1948), who showed that the higher the number of partners, the higher the fitness for
the male function becau se it produces small sperm cells. Nevertheless, female
function maximizes its fitness by seeking high quality mates because it produces
large eggs and this function has to invest in cocoon production. As a consequence,
there is a conflict between the sexes. Indeed, Porto et al. (2008) found a negative
relationship between the present investment in male function and the future fertility
of the female function in their research on E. andrei . Sexual selection is expected to
occur because of female function as long as a sufficient number of mates are
5.2.1 Precopulatory Sexual Selection
Copulation is very costly and involves sperm and mucus production and long
periods of time. Consequently, precopulatory selection is expected in environments
where the density of earthworm s is high.
5 Reproduction of Earthworms: Sexual Selection and Parthenogenesis 71
One of the factors that could influence precopulatory sexual selection is the
female fecundity of the partner, which may be related to body size. Large earth-
worms have not been found to produce more cocoons (Tato et al. 2006; Butt and
Nuutinen 1998) but they do tend to produce heavier cocoons and larger offspring
(Michiels et al. 2001). Size-assortative mating was indeed observed in the field for
the epigeic E. fetida (Monroy et al. 2005) and for the endoge ic H. elisae (Novo et al.
in press), as well as in laboratory experiments for the anecic L. terrestris (Michiels
et al. 2001). Earthworms selected similar-sized partners. Because ever y earthworm
seeks a bigger partner, equilibrium is finally reached, resulting in partners with a
similar weight, thus balancing the expectations of both mates on female and male
functions. In the particular case of epigeic and anecic worms, which can copulate at
the surface, this general tendency could be reinforced by a trade-off; worms can
either select a bigger, more fecund partner or a smaller partner, which would
decrease the risk of predation.
In ongoing laboratory experiments with H. elisae, we have observed that there is
no such size selection in virgin individuals, although the bigger virgin individuals
always managed to copulate so they seem to be more desirable.
Aside from size, reciprocation is sought from a potential partner. In simulta-
neous hermaphrodites, the primary purpose of mating is to fertilize the eggs of their
partners, rather than to fertilize their own eggs. Therefore, the conflict of two
earthworms copulating would be the amount of sperm that each of them is allowed
to give (Michiels 1998).
Finally, the quality of the place where cocoons are deposited after copulation
and the suitability of the burrow for offspring development (i.e., the moisture or
litter content) could be important factors for precopulatory assessment. Ortiz-
Ceballos and Fragoso (2006) studied parental care in Pontoscolex corethrurus
and Balanteodrilus pears ei . They found that both species build up a chamber that
they periodically clean and surround with fresh casts where a single cocoon is
deposited. Grigoropoulou et al. (2008) observed that L. terrestris deposits the
cocoons inside burrows, which may offer a protective location from the physical
environment or may represent parental investment as they were also found to be
coated with earthworm casts. These casts could be a means of maintaining the
moisture content or protecting cocoons from predators.
The mechanism through which earthworms choose a mate, assess size, test
reciprocity, or assess the burrow quality of their potential partners remains
unknown, although there are some data on these factors. Chemical cues have
been suggested in earthworms as a mechanism of finding and attracting the mate
(Olive and Clark 1978; Edwards and Bohlen 1996).
Grove and Cowley (1926) suggested the existence of a courtship in E. fetida as
they observed short and repeated touches between partners before mating. This type
of contact, executed with the prostomium, was also observed by Nuutinen and Butt
(1997)inL. terrestris and could last 90 min. The prostomium has been described as
a sensory lobe with many chemoreceptors or sensory cells (Wallwork 1983).
During contact, the clitellum and associated structures could be indicators of
female functionality and glandular margins of the male pores could be indicators
72 D.J.
az Cosı
n et al.
of male functionality. These structures could provide a means of evaluating the
partner and assuring reciprocation. Reciprocation can also be assured by increasing
the copulation time, which would prevent the partner from copulating with other
earthworms. In addition, Nuutinen and Butt (1997) observed that L. terrestris
visited the potential mate’s burrow by inserting its anterior segments, but retaining
the posterior segments in their own burrows, as a mechanism to evaluate the
In case of the size assessment, it is also suggested that assortative mating could
be due to a physical incompatibility of the copula among individuals of differ ent
sizes (Michiels et al. 2001), although this incompatibility would only result from
large differences in size.
These selective forces depend on other factors, such as the density of earth-
worms or the distance of potential mates. Indeed, the low dispersal ability of these
animals provides a restriction in the number of available mates. Earthworms have
low migration rates, with observed natural dispersal rates of only 1.4–9 m year
(Ligthart and Peek 199 7 ; Hale et al. 2005) and are therefore expected to mate with
partners living in their vicinity. In addition, in the case of the earthworms who
copulate at the surface, a smaller distance to the partner would also minimize the
risk of predation. There is evidence for this selective limitation produced by
distance. Nuutinen and Butt (1997) investigated burrow visit patterns in L. terrestris
and found that the nearer the burrow opening was, the more visits the worms made
to assess the potential part ner quality. In addition, Sahm et al. (2009) showed mate
choice in the same species for its closest partner and Novo et al. (in press) found that
H. elisae do not move long distances to find mating partners. Nevertheless, this low
dispersal could cause inbreedi ng, which is generally accepted to be unadaptative
and would reduce the fitness of the offspring. Partner selection has not been found
to be dependent on relatedness (i.e., kin recognition), and Novo et al. (in press) did
not find a correlation between mating probabilities and the level of heterozygosity
in H. elisae. Regarding this, differential investment in offspring is thought to occur
(Velando et al. 2006, see Sect. 8.2.2).
Finally, parasite concentrations may influence mate choice, since they can have
a negative effect on earthworm growth as shown by Field and Michiels (2005) for
the association between Monocystis and L. terrestris. In addition, earthworm skin
color could be positively correlated with parasite concentration (Field et al. 2003),
which could be a sign used to evaluate partners. Nevertheless, Sahm et al. (2009)
failed in an attempt to show a relationship between parasite concentration and mate
choice, and more studies are neede d to assess this correlation.
5.2.2 Postcopulatory Sexual Selection
In spite of the precopulatory sexual selection, multiple mating is common in
earthworms (Monroy et al. 2003;Sahmetal.2 009;Novoetal.inpress)andall
the allosperm receive d is stored and sometimes mixed (Novo et al. in press) in the
5 Reproduction of Earthworms: Sexual Selection and Parthenogenesis 73
spermathecae. Therefore, postcopulatory sexual selection such as sperm competi-
tion (Parker 1970) or cryptic female choice (Thornhill 1983) could be expected.
The sperm remains viable in the spermathecae until fertilization. Butt and
Nuutinen (1998) observed that L. terrestris was capable of successfully maintaining
the received sperm up to 6 months. Meyer and Bowman (1994) reported that
E. fetida cont inued cocoon production for up to 12 months after the earthworms
were isolated from their partner, although these authors did not measure viability.
n et al. (2003) reported sperma thecae full of spermatozoa during diapause in
H. elisae. This would be advantageous for species with poor dispersal capac ities or
for species that occur in low densities that can copulate at any time of the year.
The maintenance of sperm for such a long time implies the existence of some
kind of preservation mechanism. There is evidence suggesting that the spermathe-
cal epithelium actively contributes to the successful maintenance of sperm by
providing a favorable luminal environment (Grove 1925; Varuta and More 1972)
or by producing nourishing substances (Vyas and Dev 1972; Jamieson 1992; Novo
et al. (unpublished data))
A possible mechanism for postcopulatory sexual selection developed by the
recipient is sperm digestion. Richards and Fleming (1982 ) observed spermatozoal
phagocytosis by the spermathecae of the facultative parthenogenetic Dendrobaena
subrubicunda and other lumbricids. This is likely related to the removal of aging or
aberrant sperm during the months when cocoon production was minimal. Novo
et al. (unpublished data) found sperm degeneration in the central area of spermathe-
cae from H. elisae (Fig. 5.1a, b). These authors also observed sperm intrusions into
the epithel ium of spermathecae with high sperm contents, although these intrusions
seemed to occur in areas where the sperm sought more nutrients rather than
phagocytosis processes (Fig. 5.1c). Future ultrastructure studies will shed light on
these mechanisms.
Fig. 5.1 Histological preparations of the spermathecae from H. elisae. Sperm degeneration (a and
b in detail). Sperm intrusions in the epithelium of the spermathecae (c)
74 D.J.
az Cosı
n et al.
Another strategy for cryptic female choice could be the differential storage of the
received allosperm within the spermathecae. The recipient can control the storage of
sperm by increasing the complexity of these organs. Different species of earthworms
have different numbers of spermathecae (Sims and Gerard 1999), although Novo
et al. (in press) demonstrated using microsatellite markers that the four spermathe-
cae from H. elisae contained sperm from the same individuals. Grove and Cowley
(1926) observed that the transmission of sperm in E. fetida typically occurs on both
sides of the individual, whereas in L. terrestris some indi viduals were found to have
spermatophores on only one side of their body (Butt and Nuutinen 1998).
Moreover, some earthworms present different sperm loads within a single
spermathecae. This has been observed in some hormogastrids (Qiu and Bouche
1998), and in Megascolides australis, in which spermatozeugmata (i.e., sperm in
orientated bundles) were reported by Van Praagh (1995). In addition, the sper-
mathecae may include one or more diverticula that arise from the duct (Butt and
Nuutinen 1998).
Finally, the amount of sperm stored in each spermatheca could be controlled,
and this occurs for L. terrestris, which predominantly store sperm in the two
posterior spermathecae when there is no injection of allohormones (Koene et al.
2005, see later). Garvı
n et al. (2003 ) also observed that the second pair of sper-
mathecae seems to be the main recipient of spermatozoa in H. elisae. However,
Velando et al. (2008) showed that E. andrei distributes the sperm equally among the
four spermathecae.
Cryptic female choice may also be achieved through differential investmen t in
offspring. Velando et al. (2006) found that E. andrei adjusted the breeding effort to
the degree of mate relatedness, showing that inbreeding and outbreeding cause a
strong reduction of cocoon production, especially in genetic lines with high repro-
ductive rates.
Sexual selection drives the evolution of strategies that increase the chances of
fertilization for the donated sperm as a means of increasing paternity. Some of these
strategies have been observed in earthworms. Velando et al. (2008) reported a
behavior that could promote sperm competition in E. andrei, which can have a high
degree of control over their own ejaculate volume after evaluating their partners.
This species donated three times as much sperm as they did normally when mating
with a nonvirgin mate. Moreover, such increases were greater when the worms
mated with larger partners. Marin
o et al. (2006) also showed a sperm trade in
E. andrei, which adjusted the amount of sper m they release to the volume they
receive from their mating partner during copulation. In addition, the total sperm
volume they found in the spermathecae was correlated to the recipient’s body mass,
indicating that this adjustment is in accordance with the quality of the partner.
Koene et al. (2002) proposed that during mating, L. terrestris use their copula-
tory setae to pierce their partner’s skin to inject an allohormone produced by the
setal glands which manipulates the reproduct ive physiology of the partner and
damages the body wall. The injection of this substance provokes a higher uptake
of sperm, a more equal sperm distribution over the four spermathecae, and an
increase the amount of time occurring before the next mating. The damage caused
5 Reproduction of Earthworms: Sexual Selection and Parthenogenesis 75
by the injection itself could incur a considerable cost that inhibits another mating
(Koene et al. 2005).
5.3 Parthenogenesis
5.3.1 Definition
Parthenogenesis is a very wide collective concept. Historically, classical authors
addressed this concept on several occasions; although not defining the concept or
providing an experimental appro ach, authors posed hints regarding the existence of
this kind of reproduction. Although Bonnet provided experimental proof for this
kind of reproduction in aphids in 1762, it was not until 1849 that Richard Owen
coined the term. He defined parthenogenesis as “procreation without the immediate
influence of a male”. As this general concept could include several typically asexual
modes of reproduction such as fission or budding, several authors attempted to
create new definitions for this term. A century later, Suomalainen (1950) defined it
as “the development of the egg cell into a new individual without fertilization”.
Later, Beatty (1957) defined it first as “the production of an embryo from a female
gamete without the concurrence of a male gamete, and with or without eventual
development into an adult”, but modified the definition in 1967 (Beatty 1967)by
substituting “without any genetic contribution from a male gamete” for “concur-
rence of a male gamete”. In this way, Beatty extended the definition to include
special types of parthenogenesis such as gynogenesis. Nevertheless, all of these
definitions give rise to some terminological difficulties.
5.3.2 Types of Parthen ogenesis in Earthworms
Several classifications have been used to define the different types of parthenoge-
netic mechanisms. To understand earthworm cla ssification of parthenogenesis, it is
worth mentioning the classifications proposed by Thomsen (1927); Ankel (1927);
Suomalainen ( 1950) and White (1973); these are mainly based on the mode of
reproduction, sex determination, and cytology.
The system of classification proposed by Thomsen (1927) and Ankel (1927)
points out two main points: the zygoid–azygoid status of an individual and the
maintenance of the zygoid chromosome number. It includes two main categories:
generative or haploid parthenogenesis (in which chromosome reduction take s place
in the eggs, and consequently the parthenogenetic offspring have an azygoid
haploid-number of chromosomes), and somatic parthenogenesis, in which parthe-
nogenetic offspring have a zygoid–diploid or polyploid-chromosome numb er.
76 D.J.
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The difference between the two categories basically depends on the absence
(apomixis) or presence (automixis) of chromosome conjugation and reduction.
Both concepts are synonymous with White’s concepts of ameiotic and meiotic
parthenogenesis, respectively.
When considering sex determination, it is especially useful to use the classifica-
tion of parthenogenesis proposed by Suomalainen et al. (1987): arrhenotoky,
thelytoky and deut erotoky, or amphitoky (unfertilized eggs producing only male
descendants, only females, or descendants of both sexes, respective ly).
Parthenogenetic earthworms are generally automictic and thelytokous. Follow-
ing the cytological studies of Muldal ( 1952); Omodeo ( 1951, 1952, among others)
and Casellato and Rodighiero (1972), there is a premeiotic doubling of the chromo-
some number at the last oogonial divisions resu lting in endomitosis, followed by
the formation of chiasmatic bivalents and regular meiosis with the extrusion of two
polar bodies. The genetic consequences of this cytological mechanism are similar
to those of apomixis (i.e., the formation of clonal animals), as synapsis is restricted
to sister chromosomes that are exact molecu lar copies of one another. The immedi-
ate genetic conse quence of this mechanism is that heterozygosity is maintained.
Following White (1973), all bivalents are structurally homozygous and multiva-
lents are never formed. Consequently, this kind of reproduction is perfectly com-
patible with different degrees of polyploidy, especially in odd-numbered levels
(Fig. 5.2).
Only one exception to the parthenogenetic mechanism described above has been
found. Dendrobaena octaedra shows apomictic parthenogenesis: the chromosome
number is not doubled in the oogonia, the chromosome number of the oocytes is
unreduced, and there is only one equational maturation division (Suom alainen et al.
1987). For this species, Omodeo (1953) and later Gates (1972; as explained later in
this chapter) described different parthenogenetic forms with a huge degree of
morphological variation, which makes it very difficult to establish the evolutionary
relationships among them. Omodeo (1953) suggested that “it could be the result of a
breakdown of developmental canalisation in the absence of stabilizing selection”,
while White (1973) indicated that “it seems more likely that it indicates the
coexistence of numerous biotypes differing significantly from one another geneti-
cally, even if not in their visible cytology”.
Parthenogenesis is one of the main sources of morphological variability within
reproductive structures of earthworms. This variability is related to the reduction in
the investment in male structures: seminal vesicl es, testes, spermathecae, genital
setae, and prostates are reduced or even lacking; there is no sperm production (i.e.,
lack of iridescence in male funnels and sperma thecae); and sperma tophores are
lacking (in some cases they are produced but are invariably empty). In Octolasion
tyrtaeum (Muldal 1952; Jaenicke and Selander 1979), male structures are not
reduced and pseudo gamy is shown: individuals copulate to exchange spermato-
phores that are invariably empty. Thus, although spermatozoids are not necessary,
this species needs a mechanical or chemical stim ulus to trigger reproduction.
Polymorphic degradation of reproductive structures is often observed in partheno-
genetic organisms. In some species, such as Eiseniella tetraedra even hypergynous
5 Reproduction of Earthworms: Sexual Selection and Parthenogenesis 77
individuals (with an extra pair of ovaries) can be found (Jaenicke and Selander
1979). However, in other parthenogenetic earthworms such as Aporrectodea tra-
pezoides, both primary and secondary male sexual characters, such as perithecal
papillae, tubercula pubertatis, spermathecae, swo llen male porophores, and seminal
vesicles, are retained. Recent studies show that pseudogamy is not observed in this
species (Ferna
ndez et al. 2010). As discussed later, this seems to suggest very
different origins of parthenogenesis in the different species.
Parthenogenesis is not homogenously distributed in earthworms; it is only found
in lumbricids and megascolecids. It is curious that it is not found (or not known to
occur) in glossoscolecids or hormogastrids; this clearly shows that their life traits or
evolutionary histories should be completely different and that somehow partheno-
genesis and even polyploidy are not compatible or viable in this family.
Fig. 5.2 Automictic parthenogenesis: the genetic consequences of premeiotic restitution
78 D.J.
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5.3.3 Parthenogenesis and Polyploidy
Most part of the parthenogenetic earthworms are polyploids. Polyploidy ranges
from tri- to dodecaploidy. From a cytogenetical point of view, automictic biotypes
should be diploid (White 1973 ); nevertheless, in parthenogenetic lumbricids, poly-
ploidy is the most common phenomenon. This is because, as explained later, the
automictic mechanism in most lumbrici ds is premeiotic doubling, which leads to
genetic consequences similar to an apomictic mechanism, leaving levels of hetero-
zygosity unchanged from generation to generation (Suomalainen 1950). Because of
premeiotic doubling, no multivalents are formed, so pairing only occurs between
genetically identical sister chromosomes; this mechanism is compatible with odd-
numbered polyploidy, as only bivalents are formed. This is the complicated chro-
mosomal background that can give rise to different ploidy levels even within the
same species. For example, in Dendrobaena rubida, diploid, triploid, tetraploid,
hexaploid, and octoploid biotypes are known to occur, which clearly shows the
extremely high liability of the genetic system. It has been proposed that automixis
could be a step before apomixis (White 1973), which could mean that most
lumbricids could be evolving toward an apomictic partheno genesis. Polyploi dy
could be common in earthworms, as animals lacking the chromos omal determina-
tion of sex are particularly prone to this kind of repr oduction, which is the main
mechanism preventing the establishment of polyploidy in animals (White 1973).
One of the main advantages of polyploidy in parthenogenetic species is the increase
in genetic variability.
Since no study to date has elucidated the origin of parthenogenetic earthworms
(as explained later in this chapter), it is not known if parthenogenetic earthworms
may have arisen from hybridisation processes. These kinds of processes have been
found to be very common mechanisms causing asexuality (only to the extent that
parthenogenesis can be considered to be asexual reproduction) in animals and
plants (Delmotte et al. 2003). Following this assumption, polyploidy (and particu-
larly allopolyploidy) could provide important advantages, such as heterosis, to
parthenogenetic species. This strong advantage could lead the parthenogenetic
morphs to have more general purpose genotypes, allowing them to adapt to a
wider range of environmental conditions than their sexual amphimictic ancestors
(White 1973). There is much evidence that hybrid vigor could be responsible for the
success of polyploids, but there is insufficient information to determine this with
5.3.4 Genetic and Ecological Consequences of Cloning
As stated by Hugh es (1989), it is extremely difficult to define the advantages or
disadvantages of parthenogenesis, as these depend on the situation; for some groups
of animals, parthenogenesis is tremendously advantage ous, while in others it is not.
5 Reproduction of Earthworms: Sexual Selection and Parthenogenesis 79
Therefore, natural selection should control the pattern of occurrence in each group
of animals.
Using molecular tools, very different degrees of genetic variability have been
reported in different species. Both with allozyme electrophoresis and with mito-
chondrial gene sequencing, genetic variability was recorded as being high in
D. octaedra (Haimi et al. 2007; Terhivuo and Saura 1996; Cameron et al. 2008)
and Aporrectodea rosea (Terhivuo and Saura 1993; King et al. 2008), but low in
O. tyrtaeum (Jaenicke et al. 1980; Heethoff et al. 2004) and O. cyaneum (Terhivuo
and Saura 2003). In A. trapezoides, both mitochondr ial and nuclear sequences
resulted in an extremely high number of clones (Ferna
ndez et al. unpublished data.).
Judging from the number and distribution of partheno genetic earthworms, one
could expect that parthenogenesis is quite advantageous in this group. Parthenoge-
netic earthworms are widespread and very abundant, especially among peregrine
species (Blakemore 1994) such as A. rosea, A. trapezoides,orO. tyrtaeum. Hughes
(1989) pointed out the following advantages of parthenogenesis: both high levels of
heterozygosity and exceptionally fit genomes, which are maintained and inherited
by avoiding recombination and segregation; high reproductive rates, which could
potentially be doubled by avoiding the production of males (i.e., no twofold cost in
parthenogenetic reproduction); high colonizing abilities, since there is no need to
mate; high values of reproductive potential, enabling clones to quickly replace
losses; advanced polymorphism generated from selection at the level of the
genome; and the delay or prevention of senescence as somatic replicas from
undifferentiated somatic cells are generated. In reference to the last advantage,
Hughes (1989) pointed out that several clones of oligochaetes did not show any
signs of senescence after having been maintained for many generations.
5.3.5 The Species Concept in Parthenogenetic Earthworms
Parthenogenetic earthworms were wisely defined as “system atist’s nightmares” by
Blakemore (1999). The biological species criterion cannot be applied to partheno-
genetic earthworms, as each individual meets the criterion of being completely
reproductively isolated not only from the parental species, but also from every sister
clone. Several authors have attempted to resolve this problem, but an agreement has
never been reached. Mayr (1963) suggested that the best solution would be to use a
morphological criterion. Following this author (1963), a parthenogenetic species
would be the one that “results in the combination of a single species of those
asexual individuals that disp lay no greater morphological difference from each
other than from conspecific individuals or populations in related bisexual species”.
He also proposed that clones can be combined into collective species when no
essential morphological or biological differences have been observed. To complete
this criterion, the author also argued that if a parthenogenetic line originated from
an amphimictic species by an irreversible chromosomal event (such as polyploidy),
it should be considered to be a separate and sibling species, although almost no
80 D.J.
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n et al.
morphological differences could exist. This criterion has traditionally been used to
define species in parthenogenetic lumbricids, though it can be difficult to apply as
the degree of morphological variation is sometimes slight and the features defining
parthenogenetic and even amphimictic species can overlap. This is a particularly
big problem in complexes of very similar species containing both amphimictic and
parthenogenetic species such as the Aporrectodea caliginosa species” complex. In
this context, other approaches, as discussed later, could be essential not only for
properly defining parthenogenetic species, but also for determining the taxonomic
status of each form in these species complexes.
Following Gates (1974), “the species is understood to include not only the
interbreeding population, but also all recently evolved uniparental strains, clones,
or morphs that clearly are affiliated with it”. This statement is useful when inter-
mediate forms are found, but still does not solve the problem of how to resolve the
status of parthenogenetic species with unknown (or extinct) amphimictic parental
species. Another option would be to use the phylogenetic concept of species based
on molecular markers, which would provide information about the genetic diver-
gence between morphs or species. However, these tools are not so well developed in
earthworms that they could provide a good idea as to the exact amount of diver-
gence that should be used to differentiate between species. In addition, there is
evidence of different degrees of divergence among closely related species in the
different earthworms groups. The best way to define a parthenogenetic species (and
amphimictic species, particularly when dealing with complex of species) is to use
an integrative concept of species, using ecological, behavioral, morphological, and
molecular data. A species should not be given a name if its biology is not well
understood, but then, it is completely necessary to name the species . Parthenoge-
netic species are very common among the earthworms, and thus a solution needs to
be found. The ideal study would be one using all of the available approaches to
examine the same individuals so as not to incorporate any source of error or
introduce any possible mistakes when identifying species. Making comparisons
with previously published data is dangerous because different authors might have
incorrectly identified species when dealing with partheno genetic morphs or species
from a complex, in which intermediate forms are typically found. The best means of
eliminating this uncertainty is to deposit the individuals used in the experiments
into a collection.
Gates (1974) categorized parthenogenetic morphs of D. octaedra using the
presence or absence of different reproductive male structures. Gates (1974) defined
morphs lacking spermathecae, male terminalia, testes, testis sacs, or seminal vesi-
cles or those lacking several of these structures (e.g., athecal anarsenosomphic, with
or without testes). He also included two categories of intermediate morphs with an
incomplete or asymmetrical deletion of the above organs: hermaphroditic parthe-
nogenetic morphs were defined as those that had reproductive organs in a juvenile
state, while hermaphroditic morphs used biparental reproduction and were also
parthenogenetic. Unfortunately, few studies have demonstrated the existence of
these forms in every parthenogenetic species ; the knowledge about the extension
and degree of parthenogenetic morphs in parthenogenetic species is quite limited.
5 Reproduction of Earthworms: Sexual Selection and Parthenogenesis 81
This is a problem both for clarifying the taxonomy of earthworms using this type of
reproduction, and for understanding the origin of parthenogenesis in these species.
Gates (1974) and Blakemore (1999) suggeste d that parthenogenetic morphs
should be given a name only when the parental amphimictic species can be
determined. We totally agree with this statement. Nevertheless, as Blakemore
suggested, the origin of the name, regardless of whether it was based on morphs
or parthenogenetic forms, has no effect on the availability of a taxonomic name
(ICZN 1999, Article 17.3). Moreover, Gates (1972) suggested that provision of
names for all intermediate morphs of such species complexes was ridiculous.
Another limitation, as stated by Suomalainen et al. (1987), is that there are still
very few examples of taxonomic diversification beyond the species level in parthe-
nogenetic earthworms.
5.3.6 The Origin of Parthenogenetic Forms
Amphimictic ancestors of parthenogenetic forms are well known in many different
animal groups, but this is not the case for Lumbricids. Hybridization has been
proposed several times (e.g., Suomalainen et al. 1987) as a common origin of
parthenogenetic animal species such as fishes, lizards, and salamanders. Among
invertebrates, there are many examples of parthenogenetic forms originating from
Hybridization in the literature. This is the case, for example, for parthenogenetic
forms in delphacid leaf-hoppers or stick insects belonging to the genus Acanthoxyla
which were described as having two haploid genomes, one of which came from an
amphimictic parental species (Buckley et al. 2008). Suomalainen et al. (1987) also
gave some examples among invertebrates in which parthenogenesis seems to have
arisen through a single mutational event, or through multiple events. In these cases,
parthenogenesis was a polyphyletic condition within a single species as, for exam-
ple, in the psychid moth Solenobia triquetrella.
Little is known about the origin of parthenogenetic earthworms. Molecular
biology will be very useful in shedding light on this topic. Several tools can be
useful in reaching this goal. Traditionally, some studies using allozymes have been
used to check genetic variability in parthenogenetic and sexually reproducing
species that are related, such as A. trapezoides and A. caliginosa (Cobolli Sbordoni
et al. 1987). However, the information obtained using this technique was not
sufficient to evaluate hypotheses regarding the origin of parthenogenetic forms.
An appropriate first approach would be to compare phylogenies using both mito-
chondrial and nuclear genes. To determine whether parthenogenetic species origi-
nated from hybridisation, alleles could be cloned in nuclear genes to check for the
presence of different haploid genomes in diploid and, especially, polyploid parthe-
nogenetic earthworms.
As stated earlier, there is a strong variation among parthenogenetic earthworms
regarding the type of parthenogenesis that is observed; most of the species are
automictic, but at least one is apomictic. Similarly, some species are pseudogamic
82 D.J.
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n et al.
while others are not; some lack spermathecae while others have an extra pair of
ovaries. The fact that parthenogenetic mechanisms are very labile in earthworms
provides strong evidence that parthenogens could have originated in a number of
different ways. Molecular biology will allow us to better understand why partheno-
genetic earthworms have been so successful.
5.4 Conclusion
Reproduction models in earthworms are much more variable than it could seem
a priori. Although direct cross-fertilization hermaphroditism may be seen as the
most usual model, it is common to find different ones as self-fertilization or
parthenogenesis. Even within the most widespread strategy, it is possib le to find
variations, such as presence of spermatophores.
During the last years, a great research effort has been made to shed light on some
aspects of sexual selection, such as mate assessment, copulatory behavior, and sperm
competition. Nevertheless, very interesting processes as origin and maintenance of
parthenogenesis in earthworms are mainly unknown. Deeper research on both aspects
would allow us to better understand the reproductive biology of these animals.
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... Generally, the loss of sexual reproduction has been thought a dead end in evolution, leading to early extinction [60,61]. But in contrast to this perception, parthenogenesis has been observed in many species of earthworm (Lumbricidae and Megascolecidae) and parthenogenesis in earthworms is often related to polyploidy or aneuploidy [62][63][64][65][66][67][68][69][70]. Muldal (1952) points out that parthenogenesis is important as it makes the retention of polyploidy possible, and also favors the spread of polyploid forms into new areas, since even a single parthenogenetic individual may establish a population. ...
... Muldal (1952) points out that parthenogenesis is important as it makes the retention of polyploidy possible, and also favors the spread of polyploid forms into new areas, since even a single parthenogenetic individual may establish a population. The association between parthenogenesis and high polyploidy in earthworms produces an unexpected level of heterozygosity, an advantageous condition that provides resistance to environmental stress [65,69]. ...
... Parthenogenesis often results in polymorphism in earthworms [67], with morphological variability mainly relating to the reduction of reproductive structures such as seminal vesicles, spermathecae, prostates, and an empty seminal chamber. [69]. In the case of A. triastriatus, lineage A with a thin and lustreless seminal chamber and no prostate gland observed was almost degenerated to parthenogenesis, while lineage B with a plump and glossy seminal chamber and small prostate glands had a tendency to parthenogenetic reproduction. ...
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Amynthas triastriatus (Oligochaete: Megascolecidae) is a widely distributed endemic species in Southern China. To shed light on the population genetic diversity and to elucidate the population differentiation and dispersal of A. triastriatus, a population genetic structure study was undertaken based on samples from 35 locations collected from 2010 to 2016. Two exclusive lineages within A. triastriatus—lineage A and lineage B—were revealed. Lineage A was mainly distributed at high altitudes while lineage B was mainly distributed at low altitudes in Southeast China. The genetic diversity indices indicated that the populations of A. triastriatus had a strong genetic structure and distinct dispersal histories underlying the haplogroups observed in this study. Combined with morphological differences, these results indicated a new cryptic subspecies of A. triastriatus. Lineage A was almost degenerated to parthenogenesis and lineage B had a trend to parthenogenesis, which suggested that parthenogenesis could be an internal factor that influenced the differentiation and dispersal of A. triastriatus. The divergence time estimates showed that A. triastriatus originated around Guangxi and Guangdong provinces and generated into two main lineages 2.97 Ma (95%: 2.17–3.15 Ma) at the time of Quaternary glaciation (2.58 Ma), which suggested that the Quaternary glaciation may have been one of main factors that promoted the colonization of A. triastriatus.
... However, earthworms exhibit parthenogenesis and are mainly concentrated in the Megascolecidae and Lumbricidae families [3]. The combination of parthenogenetic strategies and chromosomal polyploidy greatly increases the heterozygosity level of earthworm taxa, which is beneficial for resisting environmental stress and adapting to a wide range of extreme environments [35]. Parthenogenesis usually leads to polymorphism in earthworms, with morphological variability related mainly to the reduction of reproductive structures, such as spermathecae, prostates, seminal vesicles, and an empty seminal chamber [35][36][37][38][39][40][41][42][43][44][45][46][47]. ...
... The combination of parthenogenetic strategies and chromosomal polyploidy greatly increases the heterozygosity level of earthworm taxa, which is beneficial for resisting environmental stress and adapting to a wide range of extreme environments [35]. Parthenogenesis usually leads to polymorphism in earthworms, with morphological variability related mainly to the reduction of reproductive structures, such as spermathecae, prostates, seminal vesicles, and an empty seminal chamber [35][36][37][38][39][40][41][42][43][44][45][46][47]. M. remanens has a tendency toward parthenogenesis, with a lack of male pores, degeneration of prostate glands, and loss of seminal vesicles. ...
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Metaphire remanens sp. nov. is widely distributed throughout Hunan Province, China. We sequenced the mitochondrial DNA to investigate its population genetic structure and genetic diversity, including cytochrome c oxidase subunit I, cytochrome c oxidase subunit II, 12S ribosomal (r)RNA, 16S rRNA, and nicotinamide adenine dinucleotide dehydrogenase subunit 1, derived from 39 individuals from seven geographic locations in Hunan Province. The genetic diversity indices showed that populations of M. remanens have a strong genetic structure and obvious dispersal histories. M. remanens did not experience population expansion, except in Xiangtan City. This may be because of its evolution toward parthenogenesis. The divergence time estimates indicated that M. remanens originated at 19.2055 Ma and then generated two main lineages at 1.7334 Ma (Quaternary glaciation). These results indicate that glaciation, geographic isolation, and dispersal ability are significant factors that influence the differentiation and dispersal of M. remanens. In this study, we describe Metaphire remanens sp. nov. in morphology.
... Written by J. J. Austrian and illustrated by Mike Curato, Worm Loves Worm uses the (in)human to qualify the biological plasticity of the worm (as it pertains to biological sex). Indeed, the earthworm is a "simultaneous hermaphrodite" (Cosín et al., 2011) as it carries both sex organs. Although Worm Loves Worm starts with the declarative "Let's be married," the text's larger purpose is not to constrict what queer partnership can be (in terms of marriage) but rather to reorient the pairing to the necessities of the ritual itself (e.g., wedding rings). ...
... It is generally accepted that parthenogenetic earthworms are automictic (i.e. mode of parthenogenesis that retains meiosis) with a premeiotic doubling of the chromosome number, followed by regular meiosis which restore diploidy in the following egg (Diaz Cosin et al. 2011). Because replicated chromosomes pair prior to meiosis I, the offspring are genetically identical to the mother (such as in apomictic parthenogenesis with suppression of meiosis), and heterozygosity is maintained (Simon et al. 2003;Lutes et al. 2010). ...
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Pontoscolex corethrurus is a well-known invasive earthworm in tropical zone which is believed to have originated from the Guayana Shield in South America and was described as parthenogenetic. A recent phylogenetic study revealed four cryptic species in the P. corethrurus complex (L1, L2, L3 and L4), among them L1 was particularly widespread and was proposed as P. corethrurus sensu stricto. Here, our aims were to investigate the genetic variation of P. corethrurus L1 in its presumed native and introduced ranges and to examine its reproductive strategy. An extensive dataset of 478 cytochrome oxidase I gene (COI) sequences, obtained in specimens sampled all around the world, revealed a weak COI haplotype diversity with one major haplotype (H1) present in 76% of the specimens. Analyses of the genetic variation of 12 L1 populations were done using both nuclear (226 AFLP profiles) and mitochondrial (269 COI sequences) genetic information. The high AFLP genotype diversity at the worldwide scale and the fact that no genotype was shared among populations, allowed to reject the ‘super-clone’ invasion hypothesis. Moreover, a similar level of mean genetic diversity indices were observed between the introduced and native ranges, a pattern explained by a history of multiple introductions of specimens from different parts of the world. At last, occurrence of identical AFLPs genotypes (i.e. clones) in several population confirmed asexual reproduction, but recombination was also revealed by gametic equilibrium analysis in some populations suggesting that P. corethrurus L1 may have a mixed reproductive strategy.
... Based on our investigation, in some localities no epigeic Drawida specimens with well-developed clitella existed. It is possible that parthenogenetic earthworms can be found among black-coloured specimens as for some representatives of other families (Blakemore 2003;Diaz-Cosin et al. 2011). In order to support this point, detailed morphological studies of many specimens of these earthworms are required. ...
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In the Sikhote-Alin and Changbai Mountains of the Amur River region, earthworms of genus Drawida inhabit the northern boundary of their natural habitat. They are represented by the epigeic and anecic life-forms, three steady colour morphs and eight valid species, yet the genetic lineages of 11 have yet to be described. Based on mt-COI gene fragment sequence data, epigeic and anecic earthworms are shown to differ from one another at the interspecific level. Polymorphism and genetic intraspecific diversity are an obligatory sign for a species in the refuge even at the boundary of its distribution where this species occupies new ecological niches. The original habitats of the ancient Drawida black morph were within the Paleo-Amur River basin in the Late Neogene. Today, the meadow-swampy drawidas protrude far north along the Amur River floodplain up to the border of the last freezing in this region in comparison to the forest earthworms. However, at similar northern latitudes such as the Kuril Islands and the Sakhalin, Drawida are absent, because the soil on these islands during the last Ice Age was permafrost. The black epigeic drawidas only live in the floodplain meadows. The grey and brown morphs of the anecic species live in same forest biotopes, where they inhabit different soil horizons.
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The invasive Asian earthworms, Amynthas tokioensis and A. agrestis , have been successful in entering North American forests in recent decades, with significant damage to both soils and above-ground environments. This success could be driven in part by a polyploid genetic system and parthenogenetic reproduction, often suggested as benefits for invasive species. Therefore, we assessed the genetic population structure, genetic diversity, and reproductive system of both species using morphological traits and panels of microsatellite markers. A total of 216 A. tokioensis and 196 A. agrestis from six sites in Vermont USA were analyzed. Although all worms were morphologically hermaphroditic, all the A. agrestis lacked the male pore (the structure allowing pass of sperm between individuals), and only 19% of the A. tokioensis possessed the male pore. All A. tokioensis earthworms were triploid (scored for three alleles for at least 1 locus, and usually several), and A. agrestis was a mix of triploid and diploid individuals. Notable was the high proportion (80%) of A. agrestis earthworms that were diploid at one site. There was clearly clonal reproduction, with identical seven- locus genotypes observed for earthworms from each site, with as many as 45 individuals with the identical genotype at one site. However, the earthworms were also genetically diverse, with 14 genotypes observed for A. tokioensis and 54 for A. agrestis , and with many singleton genotypes (a single individual). Most genotypes (71% for A. tokioensis and 92% for A. agrestis ) were found at a single site. The greatest number of genotypes was found at a commercial nursery where fully 23/26 A. agrestis earthworms were singleton genotypes. As expected for the pattern of private clone alleles at sites, several measures of geographic genetic differentiation were positive, and as expected for triploid systems, an AMOVA analysis showed high within-individual genetic diversity. The paradox of clear clonal reproduction, but with a great number of genotypes for each species, and the mix of triploid and diploid individuals could be explained if the worms have been sexually reproductive, with the switch to the uniparental system only recently (or even if sexual reproduction is episodic). Last, a large number of microsatellite loci were recovered for each species and there sequence and suggested PCR primers are provided for free use by other researchers.
Genetic differentiation of the amphimictic earthworm A. caliginosa populations was investigated in the territory of Ukraine by analyzing the variability of the polyallelic locus Es-4. It was established that the settlements of this species are characterized as Fst = 0.13, which means genetic heterogeneity above the average level. The values of this index depend on the size of the population groups. Moreover, these changes can be represented as a leap from insignificant interdeme differences to statistically significant ones, obtained in the analysis of geographically remote populations, with the stabilization of the Fst index values on the macroscale. This situation is reasonable for the model of a genetically homogeneous settlement that arose once over a large area, the secondary differentiation of which was caused by internal migrations and the founder effect, which is in agreement with the regularities observed for populations of this species in North America. The comparison of the spatial differentiation of A. caliginosa populations with the genetically, ecologically, and arealogically close parthenogenetic A. trapezoides species within Ukraine showed that the apomictic species has a different type of geographical differentiation of populations. It is characterized by a vicarious structure of settlements in which one clonal form replaces another and the genotypes of clones of distant populations differ the most. This means that there are no migrations between remote settlements of earthworms within Ukraine, and the reasons for the alternative nature of the genetic subdivision of amphimictic and apomictic species settlements are associated with the mechanisms of forming genetic diversity. In amphimictic species populations, the maximum genotypic diversity is achieved through recombinations and is realized at the individual level within populations, while it is due to mutations in clonal species and appears as intergroup variability.
Present study deals with cocoon biology of tropical epigeic earthworm Perionyx excavatus (Perrier), Perionyx ceylanensis (Michaelsen) and Eudrilus eugeniae (Kinberg). Ultra structure observed under Scanning Electron Microscope (SEM) revealed that wall of the cocoon is porous (which helps in aeration) and the terminal part of the cocoon is softer than that of middle part which facilitates the hatching process. The size of the cocoon varies among the species and is related to the diameter and length of the clitellum. All the epigeic species under study are continuous breeder with high fecundity. Among the studied species shortest incubation period, highest fecundity and highest hatching rate were observed in P. ceylanensis. Cocoon production in these three epigeic species increases in summer and monsoon season and decreases in the winter season. Temperature is the limiting factor for cocoon production as well as incubation period of cocoon. Cocoon production was affected by temperature and body weight of the earthworm. Thus it can be considered that cocoon production in epigeic earthworm species depends on biological factor (body mass), as well as, ecological factor (temperature). Continuous breeding strategy, high fecundity, high rate of cocoon production, as well as high hatching rate, indicates greater usefulness of P. ceylanensis followed by E. eugeniae than P. excavatus in the vermiculture based biotechnology in North-East India.
The gametogenesis of the earthworm Hormogaster elisae, an endemic species of central Spain, was studied over a 12-month period. The ovaries, seminal vesicles, male funnels, and spermathecae of 156 specimens were removed by dissection. Microscopic analysis of these organs allowed the study of the gametogenic cycle, and provided information on copulation method and reproductive cycle. Individuals of H. elisae have two pairs of spermathecae, the posterior of which is more important for the storage of sperm. In summer, the earthworms enter quiescence, and oogenesis and spermatogenesis are interrupted. The gametogenic processes occur mainly during autumn and winter. Ovules are produced during all months except August and September, and the spermathecae contain sperm over the full 12-month period. There is a reproductive peak in spring, when most ovules are produced and the clitellum is most developed.
Clone pool structure of four ecologically dissimilar parthenogenetic earthworms (Lumbricidae) was studied by means of enzyme electrophoresis in Sweden, Finland and Estonia. Each Octolasion cyaneum population had only one clone (= electrophoretic overall phenotype) but the clones differed from each other. Separate populations probably originate from one or few founder worms or cocoons unintentionally introduced by man. In Aporrectodea rosea the clonal heterogeneity was high. This is probably due to a high rate of unintentional introductions by man and/or the existence of amphigonous populations or individuals in addition to parthenogenetic ones. Sweden and Estonia share 38% of the A. rosea clones and 49% of the A. rosea individuals with the same genotype whereas the corresponding figures between Finland and Estonia and between Finland and Sweden were lower. There is evidently some clone flow between the countries. These inter-regional clones are most probably southern immigrants into the area. The clone pool diversity of two epigeics namely Eiseniella tetraedra and Dendrobaena octaedra was high but clone pool similarities between the countries were, on average, lower than they were in A. rosea. In E. tetraedra the highest similarity was found between Finland and Sweden but in D. octaedra between Sweden and Estonia as well as Finland and Estonia, These differences can be explained by the different post-glacial dispersal patterns of the clones. Intraspecific differences in somatic traits and secondary reproductive organs between the countries were found, too. The study shows that parthenogenetic earthworms with a continuous geographical range by the Baltic Sea actually comprise populations which differ considerably in their genetical make-up and morphological and morphometric traits. The differences can best be explained by dissimilar post-glacial immigration and adaptation.