Distribution and Cytogenetic Features of Triploid Males of Crucian Carp in Azov Basin

When studying uni-bisexual crucian carp (Carassius auratus gibelio) populations in the Azov basin in 1995–2000, we found triploid males, which constituted 2.5%, on average, of the total numbers of studied samples. The areas of nuclei of erythrocytes of triploid males were, on average, 1.35 times those in diploid males. At the same optical density of DNA, the sizes of mature spermatozoon heads in triploid males were, on average, 1.8 times smaller than in diploid males, as follows from the data obtained in summer 1996. The results of similar studies carried out during the period of natural spawning activity in 1997–1999 suggest that the sizes of spermatozoon heads in triploid males were, on the contrary, 1.5 those in diploid males. Triploid males were characterized by mosaicism of spermatozoon sizes and chromosome mosaicism in somatic cells. Electrophoretic analysis for the locus of transferrin confirmed the triploid status of this genetic group. The results of comparative crosses of crucian carps with different ploidy suggest a high fertilizing capacity of triploid males, as well as normal viability of their progenies. A distinct positive correlation (r = 0.73) was found between the numbers of triploid females and triploid males in mixed di-triploid populations. No significant correlation was found between males and females within di- and triploid forms.     

Sexual maturation in triploid rainbow trout, Salmo gairdneri Richardson

This paper compares some morphological and endocrinological characteristics of diploid and triploid rainbow trout.
Significant differences were found between diploid and triploid females in GSI, condition factor, gut weight, liver weight and percentage dress-out, and between diploid and triploid males in GSI, condition factor and gut weight.
Diploid females had large, well-developed ovaries containing yolk-filled secondary oocytes whereas the triploids had only string-like ovaries containing nests of oogonia. No primary oocytes were present.
All the diploid males produced copious quantities of milt but it was possible to express a thin, watery milt containing motile spermatozoa from only two of the 12 triploid males. Testes weights in triploids were similar to those of diploids but, while the diploid testes were packed with spermatozoa, those of the triploids consisted mainly of spermatocytes and spermatids with few spermatozoa present. Measurements of the heads of spermatozoa revealed that those from triploids were larger and had a wider size range than those from diploids.
Levels of testosterone and 11-ketotestosterone in triploid and diploid males were not significantly different. However, levels of testosterone and 17β-oestradiol in diploid females were considerably higher than those of triploid females.     

Diploid males and their triploid offspring in the paper wasp Polistes dominulus

Although the hymenopteran sex-determining mechanism generally results in haploid males and diploid females, diploid males can be produced via homozygosity at the sex-determining locus. Diploid males have low fitness because they are effectively sterile or produce presumably sterile triploid offspring. Previously, triploid females were observed in three species of North American Polistes paper wasps, and this was interpreted as indirect evidence of diploid males. Here we report what is, to our knowledge, the first direct evidence: four of five early male-producing Polistes dominulus nests from three populations contained diploid males. Because haploid males were also found, however, the adaptive value of early males cannot be ignored. Using genetic and morphological data from triploid females, we also present evidence that both diploid males and triploid females remain undetected throughout the colony cycle. Consequently, diploid male production may result in a delayed fitness cost for two generations. This phenomenon is particularly relevant for introduced populations with few alleles at the sex-determining locus, but cannot be ignored in native populations without supporting genetic data. Future research using paper wasp populations to test theories of social evolution should explicitly consider the potential impacts of diploid males.     

Comparison of the gonadal development and plasma levels of sex steroid hormones in diploid and triploid sea bass, Dicentrarchus labrax L

The goal of this study was to compare the reproductive physiology of triploid and diploid European sea bass (Dicentrarchus labrax L.). Gonads of diploid and triploid fish (males and females) were examined both microscopically and macroscopically, together with the plasma levels of the major sex steroids produced (testosterone and estradiol-17beta) when fish were adults. Prior to sexual maturation, the gonadosomatic index (GSI) of triploid males was similar to that of diploids. However, the GSI in 4-year-old adult triploid males was 1.8 times lower than that of diploids (P < 0.05). All diploid males exhibited normal gonadal development. In contrast, in triploid males spermatogenesis was impaired during late meiosis, affecting severely spermiogenesis. This was achieved by an increasing imbalance in the amount of DNA present in daughter cells of the same type as spermatogenesis progressed, as demonstrated by abnormal cell sizes, culminating in inviable spermatids. Thus, no spermiating triploid fish were observed during 4 years, which included three full consecutive maturation cycles. Furthermore, the germ cells from triploids were significantly larger than those from diploids (P < 0.001). Seasonal profiles of plasma levels of testosterone in 4-year-old males were essentially similar in both ploidies. On the other hand, triploid females had rudimentary ovaries containing oogonia and primary oocytes that were arrested during meiotic prophase I, while diploid females exhibited all stages of ovarian development. Diploid females showed levels of testosterone and estradiol-17beta significantly higher than those of triploids (P < 0.05), in which no endocrine signs of maturation were observed at all. Regarding sex ratios, triploids had 10% more females than diploids (P < 0.05) but in both ploidies males predominated, as is usually found in this species under culture conditions. These results show that triploidy blocked the initial phases of meiosis in females and the latter ones in males, resulting in the absence of or reduced gonadal development, respectively. In conclusion, we provide an explanation for the lack of gonadal development in triploid male fish, and, to the best of our knowledge, we report for the first time a case in which induced triploidy completely blocks meiosis in both sexes, thus conferring functional sterility in the sea bass.     

Coexistence of pseudogamous and sexual planthoppers of the genus Ribautodelphax (Homoptera, Delphacidae)

ABSTRACT.

• 1 The occurrence of pseudogamous triploid females in populations of the diploid species Ribautodelphax pungens (Ribaut) was studied throughout Europe.
• 2 Considerable differences in triploid frequencies were found between populations but no regular geographic pattern was discerned.
• 3 Within populations triploid frequencies proved to be stable from generation to generation.
• 4 The twofold reproductive advantage of the pseudogamous triploid females is counterbalanced by active mate discrimination by diploid males against the pseudogamous triploid females in populations with high triploid frequencies.
• 5 Sexual diploid and pseudogamous triploid females showed no differences in phenology.
• 6 Differential winter mortality was found between diploid and triploid larvae.

     

Disruption of normal meiosis in artificial inter-populational hybrid females of <Emphasis Type="Italic">Misgurnus</Emphasis> loach

Artificial cross between two genetically different populations of Japanese Misgurnus loach was made to examine the reproductive capacity of the artificial inter-populational hybrid females. Ploidy status and microsatellite genotypes of the eggs laid by these hybrids were inferred from those determined in progenies developed by normal fertilization with haploid loach sperm, induced gynogenesis with UV-irradiated goldfish sperm and/or hybridization with intact goldfish sperm. Some hybrid females laid unreduced diploid eggs genetically identical to the mother. However, these diploid eggs could not develop by spontaneous gynogenesis, but grow to triploid by incorporation of a sperm nucleus. Other hybrid females laid haploid eggs together with diploid eggs and/or various aneuploid and polyploid eggs. Thus, a disruption of normal meiosis occurred in inter-populational hybrid females. The results suggested that the two populations should be so distant as to give rise to atypical formation of unreduced and other unusual eggs in their hybrids.     

Triploidy and mosaicism in Liolaemus chiliensis (Sauria: Tropiduridae).

A remarkable diversity of ploidy, i.e., diploidy, triploidy, and diploid-triploid mosaicism, was found among the somatic and testicular cells in a natural population of the lizard Liolaemus chiliensis from central Chile. Intra pop ulation, intersexual, and intraindividual ploidy variation is reported. In contrast with other species of polyploid reptiles, 86% of L. chiliensis males were mosaics (2n/3n) and 14% were diploids; 33% of females were triploid, 57.1% were mosaics, and 9.5% were diploid. Among 21 specimens, no triploid males were found in this sample. In the mosaic males, the diploid and triploid spermatogonia both enter meiosis, producing both reduced and unreduced metaphase II spermatocytes, most of them euploids. We discuss the origin for this ploidy in this iguanid lizard.     

Simultaneous formation of haploid, diploid and triploid eggs in diploid–triploid mosaic loaches

In the loach Misgurnus anguillicaudatus , very few diploid–triploid mosaic individuals, which are generated by accidental incorporation of the sperm nucleus into diploid eggs produced by clonal diploid loach, occur in nature. Ploidy examination of gynogenetic progeny induced by activation with ultraviolet-irradiated goldfish sperm indicated that diploid–triploid mosaic females laid haploid, diploid and triploid eggs, simultaneously. In addition, triploid eggs exhibited larger egg sizes. Microsatellite genotyping of diploid–triploid mosaics revealed that triploid genotypes of mosaic mothers possessed two alleles specific to the clonal diploid and one allele from normal diploid male. Diploid eggs from a mosaic mother had genotypes absolutely identical to the diploid clone. Most genotypes of triploid eggs were identical to the mosaic mother, and one of the three alleles of the mosaic mother was transmitted to haploid eggs. These results suggested that diploid germ cells, which had a clonal genome, were differentiated into clonal diploid eggs, and triploid and haploid eggs were produced from triploid germ cells in the same ovary of mosaic individuals.     

Life‐history traits of the Whiting polyploid line of the parasitoid Nasonia vitripennis

In hymenopterans, males are normally haploid (1n) and females diploid (2n), but individuals with divergent ploidy levels are frequently found. In species with ‘complementary sex determination’ (CSD), increasing numbers of diploid males that are often infertile or unviable arise from inbreeding, presenting a major impediment to biocontrol breeding. Non‐CSD species, which are common in some parasitoid wasp taxa, do not produce polyploids through inbreeding. Nevertheless, polyploidy also occurs in non‐CSD Hymenoptera. As a first survey on the impacts of inbreeding and polyploidy of non‐CSD species, we investigate life‐history traits of a long‐term laboratory line of the parasitoid Nasonia vitripennis (Walker) (Hymenoptera: Pteromalidae) (‘Whiting polyploid line’) in which polyploids of both sexes (diploid males, triploid females) are viable and fertile. Diploid males produce diploid sperm and virgin triploid females produce haploid and diploid eggs. We found that diploid males did not differ from haploid males with respect to body size, progeny size, mate competition, or lifespan. When diploid males were mated to many females (without accounting for mating order), the females produced a relatively high proportion of male offspring, possibly indicating that these males produce less sperm and/or have reduced sperm functionality. In triploid females, parasitization rate and fecundity were reduced and body size was slightly increased, but there was no effect on lifespan. After one generation of outbreeding, lifespan as well as parasitization rate were increased, and a body size difference was no longer apparent. This suggests that outbreeding has an effect on traits observed in an inbred polyploidy background. Overall, these results indicate some phenotypic detriments of non‐CSD polyploids that must be taken into account in breeding.     

Diploid males sire triploid daughters and sons in the parasitoid wasp Cotesia vestalis

In the Hymenoptera, males develop as haploids from unfertilized eggs and females develop as diploids from fertilized eggs. In species with complementary sex determination (CSD), however, diploid males develop from zygotes that are homozygous at a highly polymorphic sex locus or loci. We investigated mating behavior and reproduction of diploid males of the parasitoid wasp Cotesia vestalis (C. plutellae), for which we recently demonstrated CSD. We show that the behavior of diploid males of C. vestalis is similar to that of haploid males, when measured as the proportion of males that display wing fanning, and the proportion of males that mount a female. Approximately 29% of diploid males sired daughters, showing their ability to produce viable sperm that can fertilize eggs. Females mated to diploid males produced all-male offspring more frequently (71%) than females mated to haploid males (27%). Daughter-producing females that had mated to diploid males produced more male-biased sex ratios than females mated to haploid males. All daughters of diploid males were triploid and sterile. Three triploid sons were also found among the offspring of diploid males. It has been suggested that this scenario, that is, diploid males mating with females and constraining them to the production of haploid sons, has a large negative impact on population growth rate and secondary sex ratio. Selection for adaptations to reduce diploid male production in natural populations is therefore likely to be strong. We discuss different scenarios that may reduce the sex determination load in C. vestalis.     

Identification of triploid individuals and clonal lines in Carassius auratus complex using microsatellites

The Carassius auratus complex in natural populations includes diploid triploid and polyploidy individuals. Diploid individuals belong to the species Carassius auratus whereas triploid and polyploidy individuals are from the subspecies Carassius auratus gibelio. Triploid individuals are all female and reproduce clonally by gynogenesis. Therefore the Carassius auratus complex is an ideal system for studying evolution of unisexual reproduction. Identification of triploid individuals and clonal lines is the first step towards understanding of the evolution of unisexual clonal lines. We examined the ability of 10 microsatellites in identifying triploid individuals in 94 individuals from Japan and China. In 40 confirmed triploid individuals and eight confirmed diploid individuals, all triploid and diploid individuals can be identified by genotyping 10 microsatellite, and four triploid clonal lines were identified. Using the 10 microsatellites we genotyped 46 adult individuals (40 females and six males) from a natural population in China and found that all six males were diploid whereas the majority of females (36 of 40) were triploid and three triploid clonal lines were detected. In 18 diploid individuals from China, all individuals showed different genotypes, suggesting there is no diploid clonal line in diploid crucian carp. A phylogenetic analysis of 94 individuals from China and Japan showed that triploid individuals and clonal lines have originated recurrently.     

Reproductive capacity of triploid loaches obtained from Hokkaido Island, Japan

Reproductive capacity was investigated in naturally occurring triploid individuals of the loach Misgurnus anguillicaudatus collected from Memanbetsu Town, Abashiri County, Hokkaido Island, Japan. These triploids have been considered to appear by accidental incorporation of the haploid sperm genome from normal diploid into unreduced diploid eggs from the clonal lineage that usually reproduces unisexually. By fertilization with sperm from the normal male, one triploid female gave many inviable aneuploid (2.1–2.7n) and very few tetraploid progeny, whereas the other produced both diploid and triploid progeny. The results suggest that at least four different types of eggs can be formed in triploid females in this locality. In contrast, no progeny hatched when eggs of the normal female were fertilized with sperm or sperm-like cells obtained from triploid males. These gametes exhibited inactive or no motility after adding ambient water. They had larger head sizes than those of normal haploid sperm and had a short or no tail. Although their ploidy was triploid or hexaploid, a small number of haploid cells were detected in the semen by flow cytometry. Thus, triploid males were generally sterile, but they have a little potential for producing very few haploid sperm.     

A cryptic clonal line of the loach Misgurnus anguillicaudatus (Teleostei: Cobitidae) evidenced by induced gynogenesis,interspecific hybridization,microsatellite genotyping and multilocus DNA fingerprinting

In Memanbetsu town, Hokkaido island, Japan, a high frequency of natural triploid loaches Misgurnus anguillicaudatus (7.4% on average) was detected by flow cytometry for relative DNA content. Among sympatric diploid females (n=6) from a single population, we found two unique females that laid unreduced diploid eggs. They gave normal diploid progeny even after induction of gynogenesis with genetically inert UV-irradiated sperm. When fertilized with normal loach sperm, some unreduced eggs developed into triploids, but the rest into diploids. Hybridization using goldfish Carassius auratus sperm gave both normal diploid loaches and inviable allotriploid hybrids possessing the diploid loach genome and the haploid goldfish genome. Microsatellite genotyping and DNA fingerprinting demonstrated that the diploid progeny developing from the unreduced eggs were genetically identical to the mother, while the triploids had some of the paternal DNA. These results indicate that the diploid eggs reproduced unisexually as a diploid clone and in other cases developed into triploids after accidental incorporation of sperm nucleus. The presence of at least one clonal line in this area was shown by the identical DNA fingerprint detected in five out of 17 diploid loaches examined.     

Triploid bumblebees indicate a direct cost of inbreeding in fragmented populations

Hymenopteran species with single-locus complimentary sex-determination (sl-CSD) face an additional cost of inbreeding because of a loss of diversity at the sex-determining locus. Laboratory studies of a range of Hymenoptera have found that a small percentage of diploid males produce viable diploid sperm, and that if these males mate, then the resultant females produce triploid offspring that are sterile. Here, we use microsatellite markers to determine the frequency of triploid individuals of Bombus muscorum and B. jonellus in a model island system. Triploids were found in populations of both species. Observed triploid frequencies of up to 8% were detected, and estimated total frequencies peaked at 20% with respect to normal diploid workers. For both species, triploid frequency was negatively correlated with surrogates of population size, providing direct evidence for inbreeding in small populations. Populations limited to <～15 km(2) of suitable habitat were particularly likely to harbour triploids. Estimated total triploid frequencies were higher in B. muscorum than in B. jonellus, perhaps due to the greater dispersal range of the latter species. Implications for the conservation of rare social hymenopterans are discussed.     

Male-biased sex ratios in mature damselfly populations: real or artefact?

1. There is ongoing controversy about whether biased sex ratios in diploid insect populations are real or an artefact caused by different behaviours and/or different catchability of the sexes. This was tested by monitoring two field and three semi-natural populations of the damselfly Lestes sponsa. 2. Capture–mark–recapture data showed that population size estimates were about twice as large for males as for females at both field sites. Independent estimates of the sex ratios based on total numbers of males and females captured supported the male bias. 3. Males had higher recapture probabilities than females due to longer times between successive visits in females. Because the same pattern was found in the semi-natural populations, the longer intervals in females are no artefact due to their lower detectability. 4. Theoretical models show that the strong temporary emigration of females tends, if anything, to overestimate female population sizes and that the heterogeneity of recapture probabilities observed in males tends to underestimate male population sizes. Hence, behavioural differences between the sexes do not cause an artificially male-biased sex ratio. 5. Spatial data show that operational sex ratios are male biased at the pond but become female biased in the plots further away from the shoreline; however because of the decrease in densities away from the shoreline, this does not result in a global even sex ratio. 6. Spatial data, temporary emigration patterns, and independent estimates suggest strongly that the male-biased sex ratios in mature damselfly populations are real.     

Sex Determination in Bees. II. Additivity of Maleness Genes in APIS MELLIFERA

Twenty-two randomly taken morphological characters were used in order to estimate the Mahalanobis generalized distance between diploid males, diploid workers, haploid males and triploid workers. It was found that adult diploid males are metamales and triploid females are slightly masculinized. These facts indicate that the maleness genes are slightly additive.     

Influence of environmental parameters on growth pattern and population structure of Carassius auratus gibelio in Eastern Ukraine

Population structure and growth parameters of Prussian carp (Carassius auratus gibelio) were studied in 12 freshwater ecosystems of the Donbass region (Eastern Ukraine). These ecosystems differed significantly with respect to their surface area, water transparency and annual concentrations of phosphorus and nitrogen. Amongst the studied ecosystems, diploid males and females as well as a smaller percentage of triploid females were found. The population structure of C. a. gibelio varied significantly in terms of the percentage of triploids and sex ratio amongst diploids. A considerable proportion of triploid females (>10%) was found in four ecosystems with intermediate surface area (38–50 ha) and relatively high growth rate of specimens. The sex ratio amongst diploids was significantly female-biased in seven of 12 ecosystems, including those where triploids were present in considerable numbers. The growth parameters of Prussian carp were significantly correlated with ecosystem characteristics, but the growth patterns of diploids and triploids were not significantly different from each other. The restricted distribution of triploid specimens suggests that the triploid form might counter diverse environmental challenges, whereas the diploid form of Prussian carp seems to be efficiently adapted to a wide range of ecosystem conditions.     

Diploids v . triploids of Rutilus alburnoides: spatial segregation and morphological differences

Rutilus alburnoides complex is a common and widely distributed Iberian cyprinid, whose natural populations include mainly diploid and triploid forms. The Guadiana populations of R. alburnoides were studied to determine whether habitat segregation and morphological differences exist between these forms. The ploidy level of each specimen was determined by measuring erythrocyte DNA content using flow cytometry. Evidence of spatial segregation between diploid males and the two female forms was found. Diploid males were best represented in the River Degebe, which was shallow, with higher temperatures (especially during the spring and summer), and silt and sandy substrate. Diploid females were found in deeper water, on steeper gradients and coarse substrata, while triploid females preferred higher current velocity and a high proportion of instream cover, especially during the spring. The ecological differences may reduce competitive interactions, and should promote a stable coexistence of the different forms. Morphological distinction between fish of different ploidy levels was not established, but differences were found between the males and females. Discriminant analysis allowed, with a 10% error, the separation of both sexes through six morphological characteristics that could be recorded in the field.     

Cytogenetic studies of Poecilia (Pisces)

Natural populations of triploid females resembling the gynogenetic teleost, Poecilia formosa (Girard), occur in northeastern Mexico where they intermingle with diploid populations of this species and the members of congeneric bisexual species such as P. mexicana or P. latipinna. Mitotic configurations from gill epithelial cells show 46 chromosomes for the diploid fishes, but 69 chromosomes for members of the triploid clones associated with P. formosa. Triploid females have erythrocytes that are significantly larger than those from diploid specimens and also show a roughly 50% elevation in the average DNA content of their somatic nuclei. Similar analyses of two functionally incompetent males of P. formosa, of a number of bisexual F₁ and F₂ hybrid offpsring from P. latipinna x P. mexicana, and of females from several other poeciliid species consistently show only diploid DNA levels and somatic chromosome complements where 22N=46. Demonstration of cytogenetic criteria by which females from triploid clones may be clearly distinguished from sympatric diploid specimens of P. formosa or P. mexicana leaves unresolved, for the present, problems of an appropriate systematic designation for natural populations of triploid gynogenetic fishes. The role of sympatric speciation in the evolution of poeciliid genomes is discussed in terms of alternative mechanisms to account for the persistence in nature of a vertebrate triploid of hybrid origin.This work was supported by grants from the National Science Foundation (GB 7393) and from the U.S. Public Health Service (GM 14644).Recipient of a Research Career Development Award from the U.S. Public Health Service (1 K3 GM 3455).     

Viability of diploid males in the parasitic wasp,Diadromus pulchellus (Hym.: Ichneumonidae)

In the HymenopteraDiadromus pulchellus diploid males have been observed in samples collected in the wild and bred in the laboratory. Using a yellow body color mutant strain, a protocol of crosses, involving inbred individuals, allows the routine production of such diploid males. These males result from the development of fertilized ova and emerge normally. Their preimaginal viability is similar to those of other individuals, and their imaginal viability does not differ from that of haploid males. Diploid males present normal external morphology and neither mosaicism nor intersexuality was observed. However, they are bigger than haploid males and have head size and wing length similar to those of females. The significance of diploid male viability in hymenopteran populations is discussed.