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GENETICS
OF SOLITARY AND GREGARIOUS EMERGENCE IN
THE PARASITOID WASP MUSCIDIFURAX RAPTORELLUS: PATERNAL
MODIFICATION OF LARVAL AGGRESSION. Richard Stouthamer1, and E.
Fred Legner2 (Contacts) ------------------------------------------------------------------------------- 1Department
of Entomology, Wageningen Agricultural University P.O.
Box 8031, 6700EH Wageningen, The Netherlands 2 Department of Entomology, University of California,
Riverside, CA 92521, USA (e.legner@ucr.edu) Summary
The pteromalid
parasitoid Muscidifurax raptorellus Kogan & Legner is
polymorphic for its oviposition behavior, fly pupae parasitized by the
solitary form always give rise to one offspring, while fly pupae parasitized
by the gregarious form give rise to more than one offspring in approximately
60% of the hosts. The inheritance of gregarious emergence, i.e. more than one
wasp emerges per host, has been studied in great detail by Legner. He found this trait to be polygenically
controlled by 2‑19 genes.
Females carrying a higher percentage of their genome from the
gregarious form have a higher percentage of their offspring emerging
gregariously from a host. More
surprisingly he also found that the father's genetic background influenced
the rate at which his progeny emerged gregariously. Males from a gregarious line, when mated with females from a
solitary line, caused the females to start having offspring more
gregariously. The opposite effect
takes place when males from the solitary line are used. Legner's hypothesis was that the males
transferred some behavior modifying substance with their sperm to the female,
which resulted in a higher or lower level of gregarious oviposition. Here we test an alternative hypothesis in
which the level of gregarious emergence from hosts is determined by not only
the number of eggs a female oviposits in a host, but also by the level of
larval aggression. The fathers influence
on the level of gregarious emergence in this hypothesis only takes place
through his contribution to the genetic makeup of the larvae, with the larvae
with a larger fraction of their genome from the solitary line showing higher
levels of larval aggression. Under
our experimental conditions the identity of the male did not influence the
level of gregarious oviposition by a female, but it did have a significant
influence on the number of larvae that survived. The maternal behavior modification hypothesis did not apply
significantly in this case. Keywords: oviposition behavior, larval aggression, clutch size ---------------------------------------------------------------------------------------------------------------------------------------- Introduction
Species of the
pteromalid genus Muscidifurax are parasitoids of synanthropic
flies. The genus IS INDIGENOUS TO
THE AMERICAS WHERE IT SHOWS GREAT DIVERSITY (Kogan & Legner
1970). THE SPECIES TYPICALLY
SUPERPARATISE WITH 2-3 EGGS BUT ONLY
ONE WASP EMERGES PER HOST.
However, in A South American species Muscidifurax raptorellus
Kogan & Legner, two populations are known that differ in the number of
wasps emerging per housefly pupa (Legner 1987a,b). The form originating from Chile produces offspring
gregariously, i.e. from approximately 60% of the hosts more than one wasp
emerges while up to 17 wasps have been known to emerge from a single housefly
pupa (Legner 1987b). ON THE OTHER
HAND, the form originating from Peru RARELY produces more than one
offspring per host. The genetics of
level of gregarious oviposition has been studied in detail by Legner (1987,
1988a,b; 1989a,b,c; 1991a,b; 1993).
The level of gregarious emergence of the offspring of a virgin female
was found to be determined by the proportion of the genome originating from
the gregarious form (Legner 1987b).
The higher this proportion the higher the level of
gregariousness. This trait appears to
be polygenically determined and 2‑19 genes are involved in coding for
this character (Legner 1991a). More
surprisingly, the level of gregariousness in offspring emergence was not only
a function of the mothers genetic makeup but the father's genetic background
also influenced the level of gregariousness in which his offspring
emerged. Considerable amounts of data
indicated that the male actually influences the females oviposition behavior
in such a way that if the male originates from a gregarious line his mate
lays her eggs more gregariously, and female lays fewer eggs per host when she
mates with males originating from the solitary line (Legner 1987b, 1988b,
1993). High levels of variation in
these experiments suggested that other mechanisms might interplay in the
paternal influencing of maternal behavior.
A number of possible behavior modifying substances, that could be
transferred from the male to the female through the seminal fluid, have been
suggested (Legner 1987b), these include hormones and bacteria. Because males of the solitary line reduce
the gregariousness of the offspring, and males of the gregarious line
increase it one would have to assume the presence of two different
substances: one that increases the level of gregarious oviposition and
another that decreases this level. Another hypothesis
to explain level of gregarious emergence suggests that it is determined by
two factors: a) the number of eggs a female lays per host and b) the level of
aggressive interactions between the larvae emerging from the eggs. The influence of the father in this
hypothesis is purely through his genetic contribution to his offspring
(factor b), while under Legner's (1987b) original hypothesis the number of
eggs a female lays is influenced (factor a).
We discriminated between these hypotheses by mating females to males EITHER
OF the gregarious form or of the solitary form and by determining in half
of the hosts the number of eggs that were oviposited and in the other half
the number of wasps that emerged. As
the mothers in these tests we used F1 hybrid females between the gregarious
and solitary line. These females were
chosen because they are known to show both kinds of responses; i.e. when
mated to a male of the gregarious line the females produce more offspring per
host than when allowed to oviposit as virgins (Legner 1987b), and when such HYBRID
females are mated to solitary males they produce fewer offspring per host
than virgin females. Materials and Methods
Culture origin: A Peruvian/Chilean
hybrid and a Chilean strain used in these experiments were maintained on
pupae of the housefly (Musca domestica L.). The hybrid was formed by MATING LARGE
NUMBERS OF virgin Chilean females to Peruvian males FOLLOWED BY
PROPAGATION OF THE CULTURE FOR 6 GENERATIONS, AFTER WHICH the
hybrid continued to reproduce solitarily in mass culture. Experimental setup: All experiments were done at 25EC
and 24 hrs light. Hybrid females
between the solitary and gregarious lines were secured by mating solitary
females with gregarious males. Daughters
of this cross were used in the experiment, where they were randomly assigned
to one of three treatment groups. The
treatment groups consisted of mating the females to males of either (a)
gregarious or (b) solitary strain or (c) leaving them unmated. To each group respectively 16, 13 and 21
females were assigned. After 24hrs
the males were removed and each female was given 20 hosts daily for
oviposition The hosts of each day
were randomly assigned to two groups of each 10 hosts. The hosts of the first group were opened
within 48 hrs after parasitization had taken place to determine the number of
eggs laid per host. The hosts of the
second group were individually placed in a gelatin capsule (size 000) for
wasp emergence. In total the
parasitization of these wasps was followed for 5 days, thus per mother 50 hosts
were opened to determine the number of eggs laid per host and 50 hosts were
kept for emergence. The experiment was
done in such a way that the mother's group identity, i.e. whether she was
mated to a solitary male or a gregarious male, was not known to the person
who did the egg and wasp counts. We
thus avoided this knowledge from influencing the results. Analysis of variance
was performed on the data derived from the two samples of fifty hosts that
were either opened for egg counts or were left for wasp emergence for each
mother: the percentage of the hosts that had been opened containing more than
one egg, the percentage of hosts from which more than one wasp emerged, the
difference between these two percentages, the total number of eggs in the egg
sample, the total number of wasps in the wasp sample and the difference
between the total number of eggs and the total number of wasps, the total
number of parasitized hosts, i.e. those containing one or more eggs, the
total number of parasitized hosts resulting in adult offspring and the
difference between these two measurements per female. No transformation was done for the
statistical analysis of the data. To
determine the significance of the differences between means a Duncan's
Multiple range test was used with alpha =0.05. Results
The results shown in
table 1 indicate that the percentage of hosts
in which more than one egg was found did not differ significantly between females
mated with a male from the gregarious line or the solitary line. There was a significant difference however
between females mated to solitary males versus virgin females in the
proportion of the eggs that were placed gregariously on a host. The percentage of wasps that emerged
gregariously did however differ significantly between all groups with that
percentage being the highest for the offspring of the gregarious male,
intermediate for the offspring of the virgin females and the lowest for the offspring
of the solitary males. The difference between the percentage of hosts with
gregarious eggs vs gregarious wasps also differed between the three
groups. The group mated to the
gregarious males showed the smallest difference, the virgin females were intermediate
while the offspring of the solitary males showed the largest reduction in the
percentage gregariousness. There were no
significant differences between the groups in the total number of eggs per 5
day sample (table
2),
however the number of wasps that emerged from the five day sample did differ
significantly: the gregarious group had a significantly higher number of
adult progeny than the solitary or virgin group. These two groups however did
not differ from each other in this characteristic. The difference between the number of eggs per five day sample
and the number of wasps per sample differed among all groups with the
gregarious group showing the smallest reduction in numbers and the solitary
the highest. Finally there
appeared to be no difference in the number of hosts that contained eggs
between the groups nor in the number of hosts giving rise to wasps nor in the
difference between the number of hosts containing eggs versus those giving
rise to wasps (table
3). Discussion
In our experiments
the number of eggs laid per host by a hybrid female was not influenced by the
type of male TO WHICH SHE WAS
MATED. However, the genetic
background of the male did influence the percentage of hosts from which more
than one wasp emerged. Consequently,
the hypothesis that the male influences the females egg laying behavior has
to be discounted in favor of the hypothesis that larvae with a large
proportion of their genome stemming from the solitary line somehow reduce the
number that will eventually emerge from a host. The number of eggs laid per mother did not differ between the
groups, and neither did the number of hosts that has been parasitized. Therefore, the reduction in the number of
hosts from which wasps emerge gregariously was caused by a reduction during
the larval stage. Clearly, the
offspring of the solitary males had the highest mortally, followed by that of
the virgin females and the least mortality took place in the offspring of the
gregarious males. Several authors
have reported that aggressive interactions take place between the larvae of
the species M. raptor Girault & Sanders. Wylie (1971) found that larvae of M.
raptor will spend approximately the first ten hours after hatching
moving about on the host while occasionally feeding on the pupa. If a host has several eggs on it, the
larva that hatches first attacks and kills most or all of the eggs. If two larvae happen to survive the first
instar, combat will take place in the second instar. Similar observations are reported by
Podoler and Mendel for M. raptor. Legner (1987b) also reports that <2% of the hosts
parasitized by the solitary form of M. raptorellus receive more
than one egg, but never more [PERHAPS “RARELY” WOULD BE BETTER HERE]
than one wasp emerges. This could
also indicate that larval aggression occurs in this species and indeed WE
observed aggression between the larvae of the solitary form of this
species. Apparently the larval
fighting generally leads to one wasp emerging per host, and does not cause
the death of all the larvae on a host, since the number of hosts in which one
or more larvae are found is only approximately two larger than the number of
hosts from which one or more wasps emerge (table 3). [THIS LAST SENTENCE IS A BIT CONFUSING & SHOULD
BE REWRITTEN] The level of
aggressiveness between the larvae can be influenced to a maximal extent when
males of the different lines are crossed with hybrid females between these lines. The fertilized eggs result in female
larvae that have on average either 75% of their genome from the solitary line
((S & X G %)& X S %)& or 25% of the solitary line ((S& X
G%)& X G%)&. [PLEASE CHECK THIS LAST PHRASE] In the first case the larvae apparently
fight which results in a reduction in the percentage of hosts that give rise
to more than one offspring (fig. 1c).
While in the second case the larvae show less aggressive interactions
and per host, more of the eggs survive to give rise to adult wasps (fig.
1a). The offspring of the virgin
females is intermediate because all of their offspring will have on average
50% of their genome stemming from the solitary line (fig 1c.). There could also be an influence of the
father on the survival of male and female larvae in these crosses. In the case of solitary fathers the
fertilized eggs, i.e. daughters have on average 75% of the genome of the
solitary line and can be expected to fight more than their male siblings,
which have 50% of their genome from the solitary line. In the case of offspring of the gregarious
males, their daughters have only 25% of their genome coming from the solitary
line while the males have 50% of their genome coming from the solitary line. Consequently, in those cases where one
male and one female are placed in a host in case of the
"gregarious" group we would expect that the one male one female
combination would be more prevalent than in case of the "solitary"
group, while at the same time the one male group should be larger in the
gregarious line than in the solitary line. [LENGTHY LAST SENTENCE] Although indeed these trends are visible
in the data (table
4) they
are rather subtle, probably mainly because the fraction of the hosts receiving
one male and female egg is limited in these crosses. The gregarious
emergence is the result of two, possibly linked, effects: the number of eggs
a female oviposits per host, this effect is purely determined by the females
genome and secondly the level of aggressive interactions between the larvae,
this level is influenced by the genetic background of both parents. Females of the solitary line lay fewer
eggs per host than the females of the gregarious line. We suspect that larvae of the gregarious
line show hardly any aggressive behavior towards each other, while the larvae
of the solitary, line are aggressive. [PLEASE CONSIDER THAT THE GREGARIOUS
STRAIN ALSO SUPERPARASITISES. THAT
IS, MORE EGGS ARE LAID IN A HOST THAN WILL SURVIVE TO ADULTHOOD. THE CAUSE OF THE MORTALITY IN THE ONES
THAT DO NOT SURVIVE COULD BE DUE TO GENETIC INFERIORITY OR TO SOME DEGREE OF
AGGRESSIVENESS. I’VE NEVER CHECKED
INTO THIS] These two groups of genes could interact in such a way that
they would enhance each other. The
higher the percentage of the genome stemming from the gregarious line the
higher the number of hosts with more than one egg and the lower the level of
larval aggression, in contrast the lower the percentage of the gregarious
genome the lower the level of gregarious oviposition and the higher the level
of larval aggression. Therefore, one
would expect a slight s shaped curve to describe the relationship
between the percentage of the genome from the gregarious line in a virgin
mother and the percentage of the hosts giving rise to more than one
offspring. When this relationship
(Legner 1987b, 1991a) is inspected indeed such a deviation, albeit slight, is
found. The findings of Legner (1987b,
1991a) on the number of genes influencing the gregarious and solitary
oviposition should be reexamined in the light of these two groups genes being
involved. The finding that within one
species both solitary, fighting forms and gregarious, non‑fighting
forms appears to be unique to M. raptorellus. This will allow the testing of some of the
theory developed by Godfray (1987) on the evolution of solitary and
gregarious oviposition. [THERE IS ALSO A
HETEROSIS IN F-1 HYBRIDS WHICH IS NOT EXPLAINED IN THIS DISCUSSION. THE HETEROSIS IS EXPRESSED IN SEVERAL
WAYS: HIGHER NUMBERS OF OFFSPRING AND
HOSTS KILLED, ETC. ] [BACKCROSSING DATA
SUGGESTGED THAT AT LEAST EIGHT LOCI WERE ACTIVELY SEGREGATING FOR GREGARIOUS
BEHAVIOR. ONLY THE RANGE IS NOTED
ABOVE] Acknowledgement
Support for P. Strippentow
from a European Union Tempus grant is gratefully acknowledged. Conversations with Molly Hunter and Marcel
Visser greatly helped to clarify our thinking about this phenomenon. Host for these experiments were kindly
supplied by Leo Koopman, André Gidding and Frans van Aggelen. References Cited
Godfray, H. C. J.
1987. The evolution of clutch
size in parasitic wasps. Am. Nat.
129: 221‑233 Legner, E. F.
1987a. Further insights into
extranuclear influences on behavior elicited by males in the genus Muscidifurax. Proc. 52 annual conf. Cal. Mosquito
and Vector Control Association: 127‑130. Legner, E. F.
1987b. Inheritance of
gregarious and solitary oviposition in Muscidifurax raptorellus. Can. Ent. 119: 791‑808. Legner, E. F. 1988a.
Hybridization in principal parasitoids of synanthropic diptera: The
genus Muscidifurax. Hilgardia 56(4): 36pp. Legner, E. F.
1988b. Muscidifurax raptorellus
females exhibit postmating oviposition behavior typical of the male
genome. Ann. Entomol. Soc. Am. 81: 522‑527
Legner, E. F.
1989a. Wary genes and
accretive inheritance in Hymenoptera.
Ann. Entomol. Soc. Amer. 82: 245‑249. Legner, E. F.
1989b. Paternal influences in
males of Muscidifurax raptorellus. Entomophaga 34: 307‑320 Legner, E. F.
1989c. Phenotypic expression
of polygenes in Muscidifurax raptorellus, a synanthropic fly
parasitoid. Entomophaga 34: 523‑530. Legner. E. F. 1991a.
Estimations of number of active loci, dominance and heritability in polygenic
inheritance of gregarious behavior in Muscidifurax
raptorellus. Entomophaga 36: 1‑18
Legner, E. F.
1991b. Recombinant males in
the parasitic wasp Muscidifurax raptorellus. Entomophaga 36:
173‑181 Legner, E. F.
1993. Theory for quantitative
inheritance of behavior in a protelean parasitoid, Muscidifurax raptorellus
(Hymenoptera:
Pteromalidae). Eur. J.
Entomol. 90: 11-21. Podoler, H. and Z. Mendel.
1977. Analysis of solitariness
in a parasite‑host system (Muscdifurax raptor ‑ Ceratitis
capitata). Ecol. Entomol. 2: 153-160 Wylie, H. G.
1971. Observations on
intraspecific larval competition in three hymenopterous parasites of fly
puparia. Can. Ent. 103: 137‑142 -------------------------------------------------------------------------------------------------------------------------------------- Table 1. Mean and standard error of the percentage of hosts that
contained more than one egg (egg/hst), from which more than one wasp emerged
(wasp/hst), and the difference between these values for hybrid females that
had mated either with a gregarious male, had remained virgin or had mated
with a solitary male. n >egg/hst >wasps/hst difference gregarious male 16 38.4
(2.39) 28.7
(2.13)
9.8 (2.93) virgin 21 33.8 (2.08) 12.3 (1.86) 21.6 (2.56) solitary
male 13 43.1 (2.65) 5.3 (2.36) 37.8 (3.25) Table 2. Mean and standard error of the total number of
offspring (eggs or wasps) and the difference between these numbers, for
hybrid females that had mated either with a gregarious male, had remained
virgin or had mated with a solitary male. n >egg/hst >wasps/hst difference gregarious male 16 60.6
(2.25) 54.8 (1.97) 5.8
(1.95) virgin 22 57.0 (1.97) 43.2
(1.72) 13.8
(1.71) solitary
male 13 58.8 (2.51) 38.7 (2.19) 20.1 (2.17) Table 3. Mean and
standard error of the number of parasitized hosts indicated by the presence of one or more eggs, by the emergence
of one or more wasps and the difference between these numbers, for hybrid
females that had mated either with a gregarious male, had remained virgin or
had mated with a solitary male. n hosts with egg host
with wasps difference gregarious male 16 41.6
(1.24) 40.3
(1.40)
1.3 (0.93) virgin 21 40.9 (1.08) 38.9 (1.22) 2.1 (0.81) solitary male 13 39.2
(1.37) 37.3
(1.55)
1.9 (1.03) Table 4. Total egg and wasps
distribution per host for females that had either mated with males from a
gregarious line, had remained virgin or had mated with males from a solitary
line. Egg allocation per host,
represented as the percentage of all hosts in egg sample containing either 1,
2, 3 or 4 eggs. Wasp distribution per
hosts represented as the percentage of hosts containing a male (M) or a
female (F) or a combination of the two. No. Sample
dissected for egg counts of mothers --------------------------------------------------------- % of hosts with 1-4 eggs
No. 1 2 3 4 hosts gregarious male 16 677 62.5 32.2 5.0 0.3 virgin 22 855 65.1 30.4 3.7 0.7 solitary male 13 507 56.8 36.9 5.3 1.0 No. Sample
left for wasp counts of ----------------------------------------------------------------- mothers % of hosts from which males and/or females
emerged No. 1M 1F 2M 2F 1M1F 1M2F 3M 3F hosts gregarious male 16 661 10.8 59.6 0 23.1 3.5
0.3 0 2.6 virgin 22 803 87.5 0 12.1 0 0 0
0.4 0 solitary male 13 483 9.3 86.1 0 3.5 1.0 0
0 0 |