|
Summary of Reviewers' Comments THIS STUDY OF A
GROUP OF HYMENOPTERA THAT DEMONSTRATES GREAT DIVERSITY IN BEHAVIOUR HAS
MERIT, BUT OMITS CRITICAL DETAILS IN "Materials & Methods" OF CULTURE
AQUISITION AND EXPERIMENTAL PROCEDURE THAT WOULD ALLOW FOR A CRITIQUE OF
PREVIOUS PUBLISHED REPORTS. AS EXPERIMENTAL ENVIRONMENT AND HOST SIZE
ESPECIALLY INFLUENCE SUPERPARASITIZATION IT IS ESSENTIAL TO CLEARLY EXPLAIN THEM. THERE IS A CRITIQUE OF ONE AUTHOR'S WORK BUT AN OMISSION OF
SEVERAL OTHERS THAT PRESENT CONFLICTING RESULTS. |
|
GENETICS OF SOLITARY AND GREGARIOUS EMERGENCE IN THE PARASITOID WASP MUSCIDIFURAX
RAPTORELLUS: PATERNAL MODIFICATION OF
LARVAL AGGRESSION. Richard Stouthamer1, and E. Fred Legner2 ------------------------------------------------------------------------------- 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
Studies with new field isolates of a Peruvian strain in 1995 by
Richard Stouthamer et al. (unpublished) have shown a greater involvement of larval
cannibalism and much complexity in these species' reproduction. Indeed survival mechanisms in parasitoids
include many behaviors; among which increased cannibalism by more aggressive
larvae may be triggered during times of host scarcity. 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. Legner has studied the inheritance of gregarious emergence,
i.e. more than one wasp emerges per host, in great detail. 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 an alternative hypothesis is tested
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 father's
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. ADD: 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 where the cultures
originated from new field acquisitions, the duration of the experiment was
shorter and the parasitization environment was different than in the earlier
investigations. 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). Typically most species
superparasitize 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). However, 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 mother’s
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 primarily 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 by factor a. These hypotheses were discriminated 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. The mothers in these tests were 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: New field collections of Muscidifurax raptorellus from Peru
and Chile were obtained in 1995. FROM WHAT SPECIFIC AREA AND
TIME OF YEAR IN EACH COUNTRY WERE THE CULTURES OBTAINED? 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. ADD
REFERENCE TO Kogan & Legner (1970). 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. WHAT ARE THE DIMENSIONS (or volume) OF THE HOUSEFLY
PUPARIA? -- WHAT IS THE PARASITIZATION
ENVIRONMENT? (e.g., In screened polystyrene vials (46 cm3),
with a basal are of 7 cm2, as in earlier experiments) 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. This avoided prior 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. CONSIDERATION
OF THE FOLLOWING SHOULD BE ADDED: Superparasitism (= insertion of more than one
parasitoid egg per host) occurs in both the Peruvian and Chilean strain, and
subsequent cannibalism by hatched larvae always follows. The Peruvian strain deposits a lower
number of eggs per host than the Chilean strain but cannibalism intensity is
higher. Therefore, it is difficult to
determine the exact number of eggs initially deposited by both species. The number of adult parasitoids that
survive is always less in the Peruvian strain and usually averages about
one. On the contrary, more adult
survivors usually occur in the Chilean strain, averaging about seven at a
host density of 20 per 24 hrs. A
standardization of host density, size, age and duration of exposure to
parasitization is essential in experiments as they influence the number of
eggs deposited and the rate of cannibalism. 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 percentages 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 or in the difference between the numbers of
hosts containing eggs versus those giving rise to wasps (table 3). Discussion
Indeed survival mechanisms in parasitoids
include many behaviors; among which increased cannibalism by more aggressive
larvae may be triggered during times of host scarcity. In the current
experiments the number of eggs laid per host by a hybrid the type of male to
which she was mated did not appreciably influence female. 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 female’s egg laying behavior has to be rather 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. Podoler
and Mendel (1977) report similar observations 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 rarely more than one wasp emerges. This could also indicate that larval
aggression occurs in this species and indeed aggression was observed between
the larvae of the solitary form of this species. Apparently the larval combat generally leads to only one wasp
emerging per host, (table 3). 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%)&. In the first case the
larvae apparently engage in combat 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 it would be expected 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. 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 determined purely by the female’s 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. It is
suspected that larvae of the gregarious line show little aggressive behavior
towards each other, while the larvae of the solitary line are aggressive. The
gregarious strain also superasitizes:
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. 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 the F-1 hybrids
that is not explained in this discussion.
The heterosis is expressed in several ways: higher numbers of offspring and hosts killed, etc. Also, backcrossing data suggest that at least
eight loci were actively segregating for gregariousness behavior. DIFFERENCES
FROM PREVIOUS STUDIES IN THE EXPERIMENTAL ENVIRONMENT, HOST SIZE, QUALITY AND
DENSITY SHOULD BE NOTED HERE TO EXPLAIN LARVAL CANIBALISM RATES. 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. Leo Koopman, André Gidding and Frans van
Aggelen kindly supplied host for these experiments.
References
Godfray,
H. C. J. 1987. The evolution of clutch size in parasitic
wasps. Am. Nat. 129: 221‑233 ADD: Kogan, M. & E. F. Legner. 1970. A biosystematic revision of the genus Muscidifurax (Hymenoptera: Pteromalidae) with descriptions
of four new species. Canad. Entomol. 102(10):
1268-1290. 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 development 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. & Z. Mendel. 1977. Analysis of solitariness in a
parasite-host system (Muscidifurax 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 |