Haploid Males Research Articles

Inbreeding depression by recessive deleterious genes affecting female fecundity of a haplo-diploid mite

Abstract The effect of inbreeding on haplo-diploid organisms has been regarded as very low, because deleterious recessive genes on hemizygous (haploid) males were immediately purged generation by generation. However, we determined such recessive genes to decrease female fecundity in a population of Schizotetranychus miscanthi Saito which is known in the Acari as a subsocial species with haplo-diploidy. In mother–son inbreeding experiments, there was no depression in egg hatchability nor in the larval survival of progeny over four generations. There was, on the other hand, significant inbreeding depression in the fecundity with increasing f-value. Crosses between two lineages, one having deleterious effects on the fecundity and the other having no such effects, established during the inbreeding, revealed heterosis, and backcrosses showed that the depression was caused by deleterious recessive(s). These results strongly suggest the existence of some deleterious genes governing only the traits of adult females in wild populations of haplo-diploid organisms.

PLOIDY DOES NOT AFFECT FLUCTUATING ASYMMETRY IN THE CARPENTER BEE (HYMENOPTERA: ANTHOPHORIDAE)

Random deviations from perfect symmetry in bilateral traits, fluctuating asymmetry, are thought to reflect developmental stability: the ability of an individual to buffer against genetic and (or) environmental stress during development (Palmer and Strobeck 1986). Fluctuating asymmetry is often regarded as an indicator of individual quality or fitness (Møller and Swaddle 1997); however, factors underlying fluctuating asymmetry are not well understood (Markow 1995). It has been suggested that heterozygosity, ploidy, and genomic coadaptation (hybridization) are genetic factors affecting the magnitude of fluctuating asymmetry (Palmer and Strobeck 1986; Markow 1995; Møller and Swaddle 1997). Insects having a haplodiploid sex-determining mechanism present opportunities to examine the relationship between ploidy and fluctuating asymmetry. In hymenopterans, females are diploid and males haploid. Thus the effects of ploidy can be investigated by contrasting males and females within a population (e.g., Clarke 1997). Diploid females, in which deleterious recessives reducing developmental stability could be masked, may be expected to have smaller asymmetry than haploid males (e.g., Brückner 1976; Smithet al.1997). In this paper, we compare the level of fluctuating asymmetry in forewing venation of males to that of females in the carpenter beeXylocopa appendiculata circumvolansSmith.

A genetic screen for zygotic embryonic lethal mutations affecting cuticular morphology in the wasp Nasonia vitripennis.

We have screened for zygotic embryonic lethal mutations affecting cuticular morphology in Nasonia vitripennis (Hymenoptera; Chalcidoidea). Our broad goal was to investigate the use of Nasonia for genetically surveying conservation and change in regulatory gene systems, as a means to understand the diversity of developmental strategies that have arisen during the course of evolution. Specifically, we aim to compare anteroposterior patterning gene functions in two long germ band insects, Nasonia and Drosophila. In Nasonia, unfertilized eggs develop as haploid males while fertilized eggs develop as diploid females, so the entire genome can be screened for recessive zygotic mutations by examining the progeny of F1 females. We describe 74 of >100 lines with embryonic cuticular mutant phenotypes, including representatives of coordinate, gap, pair-rule, segment polarity, homeotic, and Polycomb group functions, as well as mutants with novel phenotypes not directly comparable to those of known Drosophila genes. We conclude that Nasonia is a tractable experimental organism for comparative developmental genetic study. The mutants isolated here have begun to outline the extent of conservation and change in the genetic programs controlling embryonic patterning in Nasonia and Drosophila.

Evidence for female mortality in Wolbachia-mediated cytoplasmic incompatibility in haplodiploid insects: epidemiologic and evolutionary consequences.

Until now, only two Wolbachia-mediated cytoplasmic incompatibility (CI) types have been described in haplodiploid species, the first in Nasonia (Insect) and the second in Tetranychus (Acari). They both induce a male-biased sex ratio in the incompatible cross. In Nasonia, CI does not reduce fertility since incompatible eggs develop as haploid males, whereas in Tetranychus CI leads to a partial mortality of incompatible eggs, thus reducing the fertility of females. Here, we study Wolbachia infection in a Drosophila parasitoid, Leptopilina heterotoma (Hymenoptera: Figitidae). A survey of Wolbachia infection shows that all natural populations tested are totally infected. Crosses between infected males and cured females show complete incompatibility: almost no females are produced. Moreover, incompatible eggs die early during their development, unlike Nasonia. This early death allows the parasitized Drosophila larva to achieve its development and to emerge. Thus, uninfected females crossed with infected males have reduced offspring production consisting only of males. Evidence of this CI type in insects demonstrates that the difference in CI types of Nasonia and Tetranychus is not due to specific factors of insects or acari. Using theoretical models, we compare the invasion processes of different strategies of Wolbachia: CI in diploid species, the two CI types in haplodiploid species, and parthenogenesis (the classical effect in haplodiploid species). Models show that CI in haplodiploid species is less efficient than in diploid ones. However, the Leptopilina type is advantageous compared to the Nasonia type. Parthenogenesis may be more or less advantageous, depending on the infection cost and on the proportion of fertilized eggs. Finally, we can propose different processes of Wolbachia strategy evolution in haplodiploid species from Nasonia CI type to Leptopilina CI type or parthenogenesis.

EVIDENCE FOR FEMALE MORTALITY IN WOLBACHIA-MEDIATED CYTOPLASMIC INCOMPATIBILITY IN HAPLODIPLOID INSECTS: EPIDEMIOLOGIC AND EVOLUTIONARY CONSEQUENCES

Until now, only two Wolbachia-mediated cytoplasmic incompatibility (CI) types have been described in haplodiploid species, the first in Nasonia (Insect) and the second in Tetranychus (Acari). They both induce a male-biased sex ratio in the incompatible cross. In Nasonia, CI does not reduce fertility since incompatible eggs develop as haploid males, whereas in Tetranychus CI leads to a partial mortality of incompatible eggs, thus reducing the fertility of females. Here, we study Wolbachia infection in a Drosophila parasitoid, Leptopilina heterotoma (Hymenoptera: Figitidae). A survey of Wolbachia infection shows that all natural populations tested are totally infected. Crosses between infected males and cured females show complete incompatibility: almost no females are produced. Moreover, incompatible eggs die early during their development, unlike Nasonia. This early death allows the parasitized Drosophila larva to achieve its development and to emerge. Thus, uninfected females crossed with infected males have reduced offspring production consisting only of males. Evidence of this CI type in insects demonstrates that the difference in CI types of Nasonia and Tetranychus is not due to specific factors of insects or acari. Using theoretical models, we compare the invasion processes of different strategies of Wolbachia: CI in diploid species, the two CI types in haplodiploid species, and parthenogenesis (the classical effect in haplodiploid species). Models show that CI in haplodiploid species is less efficient than in diploid ones. However, the Leptopilina type is advantageous compared to the Nasonia type. Parthenogenesis may be more or less advantageous, depending on the infection cost and on the proportion of fertilized eggs. Finally, we can propose different processes of Wolbachia strategy evolution in haplodiploid species from Nasonia CI type to Leptopilina CI type or parthenogenesis.Corresponding Editor: W. T. Starmer

Absence of single-locus complementary sex determination in the braconid wasps Asobara tabida and Alysia manducator

In species with single-locus complementary sex determination (sl-CSD), sex is determined by multiple alleles at a single locus. In the haplodiploid Hymenoptera, sl-CSD results in females, if individuals are heterozygous at the sex locus, and in males, if individuals are hemizygous (haploid males) or homozygous (diploid males). Several hymenopteran species have been shown to have sl-CSD, but in several others sl-CSD is absent and the phylogenetic distribution remains unclear. In the family Braconidae, all four species tested so far were shown to possess sl-CSD. In this study, inbreeding experiments were used to test for the presence of sl-CSD in two species belonging to a subfamily of the Braconidae, Asobara tabida and Alysia manducator (Alysiinae). In both species inbreeding experiments showed no difference in brood size or sex ratio compared to the (outbred) control group. Furthermore, the sex ratios found in the inbreeding treatment differed significantly from the sex ratios expected under sl-CSD. Therefore, we conclude that sl-CSD is absent in these species. This study is the first to show the lack of sl-CSD in species of the Braconidae family and that hymenopteran sex-determining mechanisms can vary, even within a family.

Mapping of hybrid incompatibility loci in Nasonia.

According to theory, F(2) hybrid breakdown (lethality or sterility) is due to incompatibilities between interacting genes of the different species (i.e., the breaking up of coadapted gene complexes). Detection of such incompatibilities is particularly straightforward in haplodiploid species, because virgin F(1) hybrid females will produce haploid recombinant F(2) males. This feature allows for screening of the complete genome for recessive genetic incompatibilities. Crosses were performed between Nasonia vitripennis (v) and its sibling species N. giraulti (g). First, a linkage map was produced using RAPD markers. RAPD markers showed an overall bias toward vitripennis alleles, a pattern not predicted by the basic two-interactor Dobzhansky-Muller model. Recovery patterns of visible markers were consistent with those of linked RAPD markers. If particular genetic interactions between two loci are causing hybrid lethality, then those genotypes should be underrepresented or absent among adult F(2) males. Four sets of significant incompatibilities were detected by performing pairwise comparisons of markers on different chromosomes. Likely explanations for the observed patterns are maternal effect-zygotic gene incompatibilities or clustering of incompatibility loci. Due to the short generation time, advantages of haplodiploidy, and availability of markers, Nasonia promises to be a productive system for investigating the genetics of hybrid inviability.

Unusually high recombination rate detected in the sex locus region of the honey bee (Apis mellifera).

Sex determination in Hymenoptera is controlled by haplo-diploidy in which unfertilized eggs develop into fertile haploid males. A single sex determination locus with several complementary alleles was proposed for Hymenoptera [so-called complementary sex determination (CSD)]. Heterozygotes at the sex determination locus are normal, fertile females, whereas diploid zygotes that are homozygous develop into sterile males. This results in a strong heterozygote advantage, and the sex locus exhibits extreme polymorphism maintained by overdominant selection. We characterized the sex-determining region by genetic linkage and physical mapping analyses. Detailed linkage and physical mapping studies showed that the recombination rate is <44 kb/cM in the sex-determining region. Comparing genetic map distance along the linkage group III in three crosses revealed a large marker gap in the sex-determining region, suggesting that the recombination rate is high. We suggest that a "hotspot" for recombination has resulted here because of selection for combining favorable genotypes, and perhaps as a result of selection against deleterious mutations. The mapping data, based on long-range restriction mapping, suggest that the Q DNA-marker is within 20,000 bp of the sex locus, which should accelerate molecular analyses.

Identification of random amplified polymorphic DNA (RAPD) markers highly linked to sex determination in the red alga Gracilaria gracilis.

The random amplified polymorphic DNA (RAPD) technique was employed in the haplo-diploid dioecious species Gracilaria gracilis to identify sex-linked PCR markers. Sixty-nine decamer oligonucleotide primers were tested on two bulks of DNA, one from five haploid males and the other from five haploid females. One of these primers (OPD13) generated a 430-bp fragment specific to males and a 620-bp fragment specific to females. The diploid individuals (tetrasporophytes) showed the co-occurrence of these two fragments. In order to verify the linkage between the sexual phenotypes and these markers, a progeny array of 59 haploid individuals (male and female) born on a diploid individual was analysed, in all of which the two markers produced by the OPD13 primer segregated perfectly with sex.

The excision of polydnavirus sequences from the genome of the wasp Cotesia congregata (Braconidae, microgastrinae) is developmentally regulated but not strictly restricted to the ovaries in the adult.

Cotesia congregata polydnavirus (CcPDV) is essential for the successful parasitism of Manduca sexta larvae by the braconid wasp Cotesia congregata. In the absence of PDV, parasitoid eggs are encapsulated. Molecular analysis has demonstrated that polydnavirus sequences are integrated in the wasp chromosomes, and an ultrastructural analysis has shown that PDV replication occurs in the calyx region in the ovaries of the wasp. The bracovirus sequences appear to be excised from the wasp genome in the calyx cells where the virus replicates. Following excision of the virus sequences, the flanking sequences are rejoined. We analysed the production of two polydnavirus circles during wasp development and in different body parts of the adults of both sexes. Our study indicates that the excision of viral sequences is developmentally regulated, beginning in the pupal stage. In the adult wasp, excision occurs ubiquitously. However, regulation in the adult seems to occur only in diploid individuals, as no excision is detected in haploid males produced from virgin females.

Effects of a Single Gene on Worker and Male Body Mass in the Fire Ant Solenopsis invicta (Hymenoptera: Formicidae)

This study examines the effects of general-protein-9 ( Gp-9 ) genotype on the body mass of polygyne (multiple-queens per nest) Solenopsis invicta Buren workers and males. We found that Gp-9 genotype was significantly associated with variation in worker mass in field but not laboratory colonies. Moreover, triploid workers with 2 distinguishable genotypes ( Gp-9BBb and Gp-9Bbb ) weighed significantly more than diploid workers with the heterozygous genotype ( Gp-9Bb ). Our results, combined with those obtained from previous studies, indicate that Gp-9 genotype, ploidy, social form, and colony queen number affect mass of S. invicta workers. We also discovered that Gp-9 genotype significantly influenced the mass of haploid males reared in both field and laboratory environments. As a group, polygyne males were significantly lighter than monogyne males, even when Gp-9 genotype was taken into account, indicating that social environment interacts with Gp-9 genotype to influence male mass. Given that diploid males previously have been shown to be lighter than haploid males, 3 factors ( Gp-9 genotype, social form, and ploidy) are now known to affect the mass of male fire ants.

A genetic perspective on mating systems and sex ratios of parasitoid wasps

AbstractParasitoid sex ratios are influenced by mating systems, whether complete inbreeding, partial inbreeding, complete inbreeding avoidance, or production of all‐male broods by unmated females. Population genetic theory demonstrates that inbreeding is possible in haplodiploids because the purging of deleterious and lethal mutations through haploid males reduces inbreeding depression. However, this purging does not act quickly for deleterious mutations or female‐limited traits (e.g., fecundity, host searching, sex ratio). The relationship between sex ratio, inbreeding, and inbreeding depression has not been explored in depth in parasitoids. The gregarious egg parasitoid, Trichogramma pretiosum Riley, collected from Riverside, CA (USA) produced a female‐biased sex ratio of 0.24 (proportion of males). Six generations of sibling mating in the laboratory uncovered considerable inbreeding depression (∼ 20%) in fecundity and sex ratio. A population genetic study (based upon allozymes) showed the population was inbred (Fit = 0.246), which corresponds to 56.6% sib‐mating. However, average relatedness among females emerging from the same host egg was only 0.646, which is less than expected (0.75) if ovipositing females mate randomly. This lower relatedness could arise from inbreeding avoidance, multiple mating by females, or superparasitism. A review of the literature in general shows relatively low inbreeding depression in haplodiploid species, but indicates that inbreeding depression can be as high as that found in Drosophila. Finally, mating systems and inbreeding depression are thought to evolve in concert (in plants), but similar dynamic models of the joint evolution of sex ratio, mating systems, and inbreeding depression have not been developed for parasitoid wasps.

Diversity of sex-determining alleles in bracon hebetor

In many hymenopterans, sex is determined at a single polymorphic 'sex locus'. Individuals that are heterozygous at this locus develop as females whereas homozygotes and hemizygotes develop as diploid and haploid males, respectively. Diploid males are developmentally inviable or sterile, and the likelihood of diploid male production depends in large part on allelic diversity at the sex locus. We assessed sex allele diversity within and among five U.S. populations of the parasitoid wasp Bracon hebetor using a series of crosses between isofemale lines. The study included two laboratory populations originating in Wisconsin, two field populations originating in Kansas and California, and a population purchased from a commercial insectary. Given the number of isofemale lines that we established, the maximum number of alleles that we could detect per population was 10. The number of sex alleles identified within populations ranged between three or four (for the two Wisconsin populations) and nine (for the California population). Subsampling three or four alleles from each population for between-population crosses led to identification of 12 alleles. Of these, four were unique to the California population, three were unique to one other population each, and one was found in only two populations. Extrapolation of the relationships between the subsampled lines led to a total estimate of 20 alleles within our lines. The relatively high allele diversity in the field and commercial insectary populations suggests that the sex determination load is relatively low in B. hebetor, and the differences in allele profiles between populations suggest that interpopulation dispersal can increase sex allele diversity within populations.

Survival of Diploid Males in Braconsp. near hebetor (Hymenoptera: Braconidae)

In the parasitoid wasp Bracon sp. near hebetor, unfertilized eggs develop as haploid males, but fertilized eggs are diploid and can develop into males or females. It is likely that the sex of diploid individuals is determined by allelic variation at a single sex locus within a system of sex determination known as single-locus complementary sex determination (CSD). Under single-locus CSD, individuals that are heterozygous at the sex locus are female and individuals that are homozygous or hemizygous are male. In B. hebetor (Say), a closely related species in which this mechanism of sex determination was 1st discovered, diploid males are sterile and usually do not survive. In this study, the survival of diploid males was confirmed in B. sp. near hebetor by the use of phenotypic and molecular markers. Females with a recessive light body color mutation (blonde) crossed with dark-bodied males produced families in which diploid males were identified through inheritance of the dominant dark-bodied allele from the father. Polymorphic RAPD-polymerase chain reaction (PCR) markers were used to confirm inheritance of paternal alleles in diploid males. In families that included diploid males, only 44% of diploids were male, indicating that diploid male survival is lower than female survival. Diploid males also are capable of mating, but mating by diploid males constrained females to producing only haploid male progeny. In the closely related species B. hebetor, diploid males rarely survive, but in other Bracon sp. (B. serinopae[Cherian] and B. brevicornis Wesmael) diploid males survive at high rates. The differential survival rates of diploid males between closely related species of Bracon may indicate that the CSD system is not as evolutionarily conserved as has been presumed.

The chromosomes of Diprion pini and D. similis (Hymenoptera: Diprionidae): implications for karyotype evolution.

Earlier cytological investigations characterize the family Diprionidae with a modal chromosome number of seven. This study shows that Diprion pini and D. similis have 14 acrocentric chromosomes (n = 14 for haploid males and 2n = 28 for diploid females). In D. pini, rRNA genes are located on the satellite and the round short arm of the chromosome carrying this segment. We studied six populations with no evidence of chromosomal polymorphism at the species level. Our results disagree with those published previously and reopen the question of karyotype evolution. Chromosome morphology, ISH and C-banding results support the hypothesis of chromosome number doubling by centric fission and not by polyploidization, followed by the growth of short arms by means of pericentromeric DNA amplification.

Fluctuating asymmetry in vestigial and functional traits of a haplodiploid insect

If natural selection on fluctuating asymmetry (FA) is common and related to trait functionality, then vestigial traits should show elevated FA. Moreover, if FA increases with heterozygosity, then haploid males of haplodiploid taxa should exhibit higher FA than diploid females. We measured fluctuating asymmetry of functional traits (fore femora of soldier morphs and disperser morphs, and wings of dispersers) and a vestigial trait (wings of soldiers) in the eusocial haplodiploid gall thrips Oncothrips tepperi (Insecta: Thysanoptera). Wing FA, but not femur FA, was substantially and significantly higher in soldiers than in dispersers in both sexes. Patterns of intersexual variation in FA were complex: for wings, female soldiers had higher FA than male soldiers but male dispersers had higher FA than female dispersers, and for femora, males and females did not differ in FA in either morph. Our results suggest that vestigial traits exhibit higher FA because of relaxation of selection for functionality, and that haploidy does not necessarily lead to increased FA in males of haplodiploid taxa.

Fluctuating asymmetry in the honey bee, Apis mellifera: effects of ploidy and hybridization

AbstractWe present a new measure of morphological asymmetry that avoids most of the statistical problems inherent in character‐by‐character analysis of size or shape. The method is an application of Procrustes analysis, which computes best‐fitting super‐positions of configurations of landmarks to the left and right sides of a single specimen. The Procrustes method combines subtle deviations in all aspects of the landmark configuration into one net asymmetry score. Directional asymmetry is separated from fluctuating asymmetry in a simple partition of a net sum‐of‐squares, and geometrical details of either component can be inspected by traditional methods of multivariate statistical analysis of landmarks.We demonstrate this method in a comparison of wing venation asymmetry in male (haploid) and female (diploid) honey bees (Apis mellifera). In addition we investigate the effects of ploidy and inter‐subspecies hybridization on asymmetry and wing venation abnormalities, using the subspecies A. m. mellifera, A. m. carnica, and the hybrid strain “Nigra”. Results suggest that while the haploid males showed a higher frequency of wing venation abnormalities and greater total asymmetry than the diploid females, most of the asymmetry difference between males and females was in the form of directional, not fluctuating, asymmetry. Hybrid females had a higher frequency of wing venation abnormalities than females of either subspecies, but there were no significant differences in the mean level of asymmetry among females of A. m. mellifera, A. m. carnica and hybrid Nigra. Hybrid males had higher absolute frequency of wing venation abnormalities and asymmetry than males of either subspecies. However the mean frequency of venation abnormalities did not differ significantly between Nigra and A. m. carnica males, and mean asymmetries were not significantly different between Nigra and A. m. mellifera males. We discuss the relationship which is assumed to exist between developmental stability and fluctuating asymmetry in light of our result.

Characterization of a microsatellite locus in the parasitoid waspAphelinus abdominalis(Hymenoptera: Aphelinidae)

AbstractPrimers for DNA amplification using the polymerase chain reaction (PCR) were synthesized for a microsatellite locus isolated from a partial genomic library of the aphid parasitoidAphelinus abdominalis(Dalman). Screening for genetic polymorphism at this locus in two laboratory strains of this wasp revealed the presence of two alleles different in the number of (GT) and (GGC) repeats. The relative frequencies of the two alleles were not significantly different between the two strains or between diploid females and haploid males. Heterozygosity at this microsatellite locus was estimated to be 0.40 which is within the range in other hymenopterous species. Given thatA. abdominalisis a good candidate for augmentative release programmes in greenhouses against aphids, we suggest that microsatellite markers may have application in discriminating among aphelinid sibling species and strains. The markers provide a means for studying the performance and impact of selected parasitoid lines on pest dynamics in field release experiments.

Development of the components of a cytoplasmic male sterility hybrid system in rye through anther culture

Anther culture was applied as a method to develop the essential components of a cytoplasmic male sterility hybrid system in rye (Secale cereale L.). These components are the male sterile seed parent (A line), its isogenic maintainer counterpart (B line) and the restorer pollen parent (R line). Australian rye cultivars were crossed reciprocally to the cultivar ‘Luchs’ which carries the Pampa male sterile cytoplasm (cms-P). Anthers of the F1s in the cms-P cytoplasm (primary cross) and their reciprocals in the normal cytoplasm (reciprocal cross) were cultured in a modified C17 medium. Male sterile and male fertile doubled haploids were obtained from the anther culture of the F1s in the cms-P cytoplasm. Testcrosses indicated that the male sterile doubled haploids were A lines and the male fertile doubled haploids were R lines (restorers). The anther culture of genotypes in the normal cytoplasm (reciprocal cross) gave all male fertile doubled haploids. Testcrosses indicated that the male fertile doubled haploids were R lines (restorers) in the normal cytoplasm. The expected maintainer B lines were not identified because of the limited number of doubled haploids obtained from the anther culture of reciprocal crosses. Experimental single cross hybrids between male sterile and restorer male fertile doubled haploids showed high levels of heterosis. The results of this study have significant economic implications especially in the production of hybrids in several species.

Use of RAPD analyses to estimate population genetic parameters in the alfalfa leaf-cutting bee,Megachile rotundata

RAPD analyses were performed on five geographically isolated populations of Megachile rotundata. We used haploid males of the alfalfa leaf-cutting bee, M. rotundata, to overcome the limitation of the dominance of RAPD markers in the determination of population genetic parameters. Sixteen primers gave rise to 130 polymorphic and 31 monomorphic bands. The unbiased estimators calculated in this study include within- and between-population heterozygosity, nucleotide divergence, and genetic distance. The genetic diversity (H = 0.32-0.35) was found to be about 10 times that of previous estimates (H = 0.033) based on allozyme data. Contrary to the data obtained at the protein level, our results suggest that Hymenoptera do not have a lower level of genetic variability at the DNA level compared with other insect species. Regardless of the different assumptions underlying the calculation of heterozygosity, divergence, and genetic distance, all five populations showed a parallel interrelationship for the three parameters. We conclude that RAPD markers are a convenient tool to estimate population genetic variation in haploid M. rotundata and that with an adequate sample size the technique is applicable to the evaluation of divergence in diploid populations. Key words : Megachile rotundata, RAPD, heterozygosity, genetic distance, nucleotide divergence.