Synapsis in Robertsonian heterozygotes and homozygotes of Dichroplus pratensis (Melanoplinae, Acrididae) and its relationship with chiasma patterns

Dichroplus pratensis has a complex system of Robertsonian rearrangements with central-marginal distribution; marginal populations are standard telocentric. Standard bivalents show a proximal-distal chiasma pattern in both sexes. In Robertsonian individuals a redistribution of chiasmata occurs: proximal chiasmata are suppressed in fusion trivalents and bivalents which usually display a single distal chiasma per chromosome arm. In this paper we studied the synaptic patterns of homologous chromosomes at prophase I of different Robertsonian status in order to find a mechanistic explanation for the observed phenomenon of redistribution of chiasmata. Synaptonemal complexes of males with different karyotypes were analysed by transmission electron microscopy in surface-spread preparations. The study of zygotene and early pachytene nuclei revealed that in the former, pericentromeric regions are the last to synapse in Robertsonian trivalents and bivalents and normally remain asynaptic at pachytene in the case of trivalents, but complete pairing in bivalents. Telocentric (standard) bivalents usually show complete synapsis at pachytene, but different degrees of interstitial asynapsis during zygotene, suggesting that synapsis starts in opposite (centromeric and distal) ends. The sequential nature of synapsis in the three types of configuration is directly related to their patterns of chiasma localisation at diplotene-metaphase I, and strongly supports our previous idea that Rb fusions instantly produce a redistribution of chiasmata towards chromosome ends by reducing the early pairing regions (which pair first, remain paired longer and thus would have a higher probability of forming chiasmata) from four to two (independently of the heterozygous or homozygous status of the fusion). Pericentromeric regions would pair the last, thus chiasma formation is strongly reduced in these areas contrary to what occurs in telocentric bivalents.     

Orientation and segregation of Robertsonian trivalents in Dichroplus pratensis (Acrididae)

Pairing behavior, metaphase I orientation, and anaphase I segregation of centric fusion trivalents were studied in 26 single, 15 double, and 2 triple male fusion heterozygotes of the polymorphic South American melanopline grasshopper Dichroplus pratensis. They represent the seven different fusions and their combinations already described in different populations of the species. Our analysis showed the following: (1) pairing behavior is very regular in all trivalents; (2) frequencies of linear orientation was very low irrespective of the trivalent involved; (3) reorientation seems to occur frequently since frequencies of abnormal segregation and aneuploid second division cells were invariably lower than those of nonconvergent orientation; (4) aneuploidy and abnormal sperm production increases with increasing number of fusions; (5) chiasma frequency and localisation is relevant to trivalent orientation since trivalents with nonconvergent orientations showed proximal and interstitial chiasmata more frequently than convergently oriented ones. The results are in agreement with the hypothesis that these polymorphisms are old and stable, and confirm that for the maintenance of a balanced polymorphism, if this polymorphism is adaptive because of its consequences on recombination, position effects, etc., changes tending to stabilise trivalent orientation and segregation are central.     

Meiotic behavior of Robertsonian heterozygotes in populations of Dichroplus pratensis (Acrididae) with different fusion frequencies

The meiotic behavior of Robertsonian heterozygotes of Dichroplus pratensis was analyzed in order to establish the nature of the fusion polymorphisms (stable or transient) that occur in the species. The range of fusion frequencies varies widely for each fusion studied and populations with extreme frequencies exist, which could indicate a tendency for the loss or fixation of a given rearrangement. Our results revealed that no significant correlation exists between orientation (convergent and non-convergent) in PMI and MI and fusion frequency, nor between aneuploidy and fusion frequency. Thus, orientation and segregation seem to be independent of the frequencies of the fusions, which also do not appear to affect severely the fertility of heterozygotes. Our data suggest that the polymorphisms are, at least, cytogenetically stable.Both authors are affiliated to the CONICET (Argentina)     

Influence of a rare unstable B-chromosome on chiasma frequency and nonhaploid sperm production in Dichroplus pratensis (Melanoplinae,Acrididae)

Three B-containing individuals of Dichroplus pratensis were found among 300 males collected in several Argentine localities. The B-chromosome is a partially euchromatic short telocentric and it is mitotically unstable. The B-chromosome number of testis cells ranged from 0 to 4 in two males and from 0 to 3 in the remaining one; this variation was inter- as well as intrafollicular. Meiotic behaviour of the Bs was very regular, association of Bs was usually chiasmate and no lagging univalents were observed during first division. The presence of Bs increased the production rate of abnormal spermatids and this effect was more pronounced in follicles with odd numbers of Bs. Chiasma frequency and between-cell variance were also increased in B-containing cells. A possible interaction between the Bs and the polymorphic centric fusions present in the species is discussed in relation to chiasma frequency determination.     

Non-random patterns of non-disjunctional orientation in trivalents of multiple Robertsonian heterozygotes of Dichroplus pratensis (Acrididae)

Metaphase I orientation of centric fusion trivalents was studied in 24 single, 19 double and 3 triple heterozygotes of Dichroplus pratensis. Different populations of this South American melanopline grasshopper are polymorphic for seven Robertsonian fusions, and the polymorphisms seem to be stable. Several cytogenetic factors involved in the orientation and segregation of the meiotic configurations such as chromosomal length, symmetry and number and position of chiasmata, have been analysed in previous works. In this paper we study another factor that is relevant in the above respect in individuals with more than one heterozygous fusion: interaction among configurations regarding orientation.Our results indicate that, when there are two or three trivalents present in the MI cell, there is an interaction in such a way that the number of metaphases in which the two or three trivalents are non-disjunctionally oriented is always significantly higher than expected under a hypothesis of independence. However, the number of cells in which all trivalents are disjunctionally oriented does not decrease significantly, so an increase of unbalanced gametes due to this factor is not expected. The stability of the polymorphisms would thus not be affected.Both authors are affiliated with the CONICET (Argentina)     

Distribution of genetic variability in populations of two chromosomal races of Dichroplus pratensis (Melanoplinae, Acrididae) and their hybrid zone

We examined, through allozyme electrophoresis, the genetic structure of populations of the acridid grasshopper Dichroplus pratensis from two chromosomal races (Northern and Southern) and their hybrid zone in Argentina. No fixed alleles for any particular race were found, although genetic differentiation among parental races was significant (0 = 0.044, 95% CI: 0.004-0.068). Hybrid populations are genetically more similar to the Southern race (0 = 0.008, 95% CI: -0.005-0.018) than to Northern ones (0 = 0.018, 95% CI: 0.002-0.030). Differential viability or fertility of hybrids, or asymmetry in mating preferences in favour of one particular cross would cause a higher proportion of matings between hybrid individuals and those from the Southern race. This would explain the high genetic similarity between those groups, in spite of their geographical vicinity with northern race populations.     

B chromosomes and Robertsonian fusions of Dichroplus pratensis (Acrididae): intraspecific support for the centromeric drive theory

We tested the centromeric drive theory of karyotypic evolution in the grasshopper Dichroplus pratensis, which is simultaneously polymorphic for eight Robertsonian fusions and two classes of B chromosomes. A logistic regression analysis performed on 53 natural populations from Argentina revealed that B chromosomes are more probably found in populations with a higher proportion of acrocentric chromosomes, as the theory predicts. Furthermore, frequencies of B-carrying individuals are significantly negatively correlated with the mean frequency of different Robertsonian fusions per individual. No significant correlations between presence/absence or frequency of Bs, and latitude or altitude of the sampled populations, were found. We thus provide the first intraspecific evidence supporting the centromeric drive theory in relation to the establishment of B chromosomes in natural populations.     

Meiosis and the Neo- XY system of Dichroplus vittatus (Melanoplinae, Acrididae): a comparison between sexes

The origin of neo-XY sex systems in Acrididae is usually explained through an X-autosome centric fusion, and the behaviour of the neo-sex chromosomes has been solely studied in males. In this paper we analysed male and female Dichroplus vittatus. The karyotype comprises 2n = 20 chromosomes including 9 pairs of autosomes and a sex chromosome pair that includes a large metacentric neo-X and a small telocentric neo-Y. We compared the meiotic behaviour of the sex bivalent between both sexes. Mean cell autosomal chiasma frequency was low in both sexes and slightly but significantly higher in males than in females. Chiasma frequency of females increased significantly when the sex-bivalent was included. Chiasma distribution was basically distal in both sexes. Behaviour of the neo-XY pair is complex as a priori suggested by its structure, which was analysed in mitosis and meiosis of diploid and polyploid cells. During meiosis, orientation of the neo-XY is highly irregular; only 21% of the metaphase I spermatocytes show standard orientation. In the rest of cells, the alternate or simultaneous activity of an extra kinetochore in the distal end of the short arm (XL) of the neo-X, determined unusual MI orientations and a high frequency of non-disjunction and lagging of the sex-chromosomes. In females, the neo-XX bivalent had a more regular behaviour but showed 17% asynapsis in the XL arm which, in those cases orientated its distal ends towards opposite spindle poles suggesting, again, the activity of a second kinetochore. The dicentric nature and the unstable meiotic behaviour of the sex neo-chromosomes of D. vittatus suggest a recent origin of the sex determination mechanism, with presumable adaptive advantages which could compensate their potential negative heterosis. Our observations suggest that the origin of the neo-sex system was a tandem fusion of two original telocentric X-chromosomes followed by another tandem fusion with the small megameric bivalent and a further pericentric inversion of the neo-X. The remaining autosomal homolog resulted in the neo-Y chromosome. This revised version was published online in July 2006 with corrections to the Cover Date.     

Postembryonic development and food consumption of Dichroplus elongatus Giglio-Tos and Dichroplus maculipennis (Blanchard) (Orhtoptera: Acrididae: Melanoplinae) under laboratory conditions

Dichroplus maculipennis (Blanchard) and D. elongatus Giglio-Tos are two of the most important melanoplines in Argentina, both ecologically and economically. The postembryonic development and forage loss (consumption of Bromus brevis Ness + fallen material) caused by older nymphs (instars IV, V, VI) and adults of both species were studied under controlled conditions (30oC, 14L:10D, 40% RH). Five nymphal instars were recorded in D. elongatus, and six in D. maculipennis. Total nymphal development was similar in both species (D. elongatus: 32 ± 0.70 days; D. maculipennis: 34.5 ± 0.37 days). Daily consumption increased from nymphal instars to pre-reproductive adult stage. In both species, pre-reproductive females had higher consumption rates than other stages considered (D. elongatus: 30.6 ± 0.56 mg dry weight/day; D. maculipennis: 48.7 ± 0.74 mg dry weight/day). In the reproductive stage, consumption decreased significantly in both sexes. When feeding, D. maculipennis let some plant material to drop, increasing total loss. The percentage of fallen material was greater in reproductive adults, representing 3.9% and 2.9% of the total daily loss for males and females, respectively. Females and males of D. maculipennis were heavier than those of D. elongatus (P < 0.05), and daily consumption was significantly higher (P < 0.05). Regardless sex and reproductive status, adults of D. maculipennis consumed 29.1 ± 0.64 mg dry weight/day on average, while one of D. elongatus 20.0 ± 0.3 mg dry weight/ day.     

FISH detection of ribosomal cistrons and assortment-distortion for X and B chromosomes in Dichroplus pratensis (Acrididae)

Assortment-distortion with respect to the X and NOR activity of a rare mitotically stable B chromosome (B(N)), was examined in 16 males of Dichroplus pratensis (Acrididae: Melanoplinae) from Argentine populations. In 1B individuals, the X and B associate preferentially during prophase I reaching a maximum level of association at zygotene. Frequency of X/B association remains relatively high up to diplotene-diakinesis and decreases steeply towards metaphase I. The percent X/B association at each stage is positively influenced by association at the previous stage, and interindividual variability in X/B association decreases as the frequency of association increases. Both chromosomes tended to preferentially orientate toward the same pole at MI (mean ratio of 16 individuals, 1.50:1) which determined an excess of XB and 00 second spermatocytes over X0 and 0B ones (1.39:1). No significant differences occurred between the MI, AI and MII assortment ratios. Fluorescent in situ hybridisation (FISH) confirmed that the B chromosome carries ribosomal genes and helped to establish that, during spermiogenesis, both the B and the normal NOR-bearing chromosome (S8) are clustered near the centriole adjunct region of spermatids. However, FISH failed to reveal the existence of inactive ribosomal cistrons in the X chromosome, as previously suggested, thus providing no support to a simple origin of the B from the X.     

Use of entomopathogenic fungi combined with biorational insecticides to control Dichroplus maculipennis (Orthoptera: Acrididae: Melanoplinae) under semi-field conditions

Grasshoppers are among the invertebrate herbivores that cause most economic losses in grasslands throughout Argentina’s Pampas and parts of Patagonia. Chemical insecticides remain the sole option for grasshopper control in this area, despite being of significant environmental concern. Our aim was to evaluate the efficacy of combinations between three entomopathogenic fungi strains (Beauveria bassiana LPSc 1067 and LPSc1082), and Metarhizium anisopliae (LPSc 907), two biorational insecticides (luphenuron and methoxyfenozide), and a new synthetic chemical pesticide (rynaxypyr) in the control of the pest grasshopper Dichroplus maculipennis under field cage conditions. Fungal strains used were adjusted to 1 × 10⁸, 1 × 10⁶ and 1 × 10⁴ conidia/ml. Insecticides were tested at three concentrations: the average concentration recommended for application in the field (100%), 50% of that level and finally 25%. Combinations of the insecticides with B. bassiana (LPSc 1067, LPSc 1082) and M. anisopliae (LPSc 907) caused higher mortality to D. maculipennis nymphs than any of the individual agents used alone. The three insecticides tested did not negatively affect the isolates of the two species of entomopathogenic fungi employed.     

Efficiency of food utilization by Dichroplus maculipennis (Orthoptera: Acrididae: Melanoplinae) on four crop plants under controlled conditions

The aim of this study was to analyze the food utilization efficiencies and the relative growth and consumption rates of different developmental stages and sexes of D. maculipennis under controlled conditions on wheat, oat, corn, and soybean plants, important crops in the Pampas region of Argentina. As expected from a polyphagous species, D. maculipennis was observed to consume all four of the plant species offered. Nevertheless, the consumption of both nymphs and adults was differentiated. Oat and wheat were more consumed than corn and soybean. Females presented higher consumption rates (384.6 ± 30.64 mg/individual/day) than males (278.71 ± 24.26 mg/individual/day). Adult females had the highest growth rate, followed by nymphs of the same sex, and then adult males. The highest values of ECI and ECD were obtained in soybean; females had higher values of food efficiencies than males, and nymphs had greater values than adults. In relation to this, soybean was the highest quality food; the amount of nitrogen present in soybean was approximately twice that found in the other species. The nutritional needs of D. maculipennis might have been satisfied by feeding on low quantities of soybean, which is, among the food offered, the most “nutritionally balanced food”.     

Infectivity,viability and effects of Paranosema locustae (Microsporidia) on juveniles of Dichroplus maculipennis (Orthoptera: Acrididae: Melanoplinae) under laboratory conditions

Infectivity and effects on host of a long-term stored aqueous suspension of Paranosema locustae on juveniles of Dichroplus maculipennis, a pest grasshopper in parts of the Pampas and Patagonia, were evaluated. Infections developed in 90–97.8% of treated individuals. Mortality increased with time, reaching highest values at 30–40 days post-inoculation (79.5–100%). Infected nymphs showed significantly slower development.     

Morphological, genetic, and chromosomal variation at a small spatial scale within a mosaic hybrid zone of the grasshopper Dichroplus pratensis Bruner (Acrididae)

Hybrid zones are regions where genetically different populations meet and mate, resulting in offspring of mixed characteristics. In organisms with limited dispersal, such as melanopline grasshoppers, hybrid zones can occur at small spatial scales (i.e., <500 m). We assessed levels of morphological, chromosomal, and molecular variability in adult males of the grasshopper Dichroplus pratensis Bruner (N = 137 males, 188 females) collected at 12 sites within a mosaic hybrid zone in a heterogeneous environment in Sierra de la Ventana, Argentina. In this hybrid zone, 2 Robertsonian chromosomal races, polymorphic for different centric fusions, meet (the "Northern race" at low altitudes and the "Southern race" at higher altitudes), forming hybrids that show monobrachial homologies during meiosis. High morphometric variation in 6 traits was revealed among grasshoppers of both sexes, with male body size positively and significantly correlated with increasing altitude. Frequency of Robertsonian fusions characteristic of the Southern race increased significantly with altitude. Moreover, fusion frequencies covaried between samples. Considerable genetic variation was revealed by random amplification of polymorphic DNA markers, with heterozygosity ranging from 0.3477 to 0.3745. Insects from low-altitude and high-altitude populations showed significant genetic differentiation, as indicated by F(ST) values. The proposed model for D. pratensis, involving the generation and maintenance by chromosomal fusions, of gene complexes adaptive in different environments, could explain the observed clinal patterns within the contact zone.     

Metaphase I orientation of Robertsonian trivalents in the water-hyacinth grasshopper, Cornops aquaticum (Acrididae, Orthoptera)

Trivalents resulting from polymorphic Robertsonian rearrangements must have a regular orientation in metaphase I if the polymorphisms are to be maintained. It has been argued that redistribution of proximal and interstitial chiasmata to more distal positions is necessary for a convergent orientation, the only one that produces viable gametes. Cornops aquaticum is a South-American grasshopper that lives and feeds on water-hyacinths, and has three polymorphic Robertsonian rearrangements in its southernmost distribution area in Central Argentina and Uruguay. The orientation of trivalents in metaphase I, the formation of abnormal spermatids and the frequency and position of chiasmata in the trivalents, was analysed in a polymorphic population of C. aquaticus. In this study we observed a correlation between the number of trivalents with the frequency of abnormal spermatids; additionally, the number of chiasmata, especially proximal and interstitial ones, was strongly correlated with the frequency of the linear orientation. Therefore we confirmed our previous assumption, based on other evidence, that the chiasmata redistribution in fusion carriers is essential to the maintenance of the polymorphisms.     

Survival and fecundity of Dichroplus maculipennis and Ronderosia bergi (Orthoptera: Acrididae: Melanoplinae) following infection by Beauveria bassiana (Ascomycota: Hypocreales) under laboratory conditions

This study examined the effects of strain Beauveria bassiana (LPSC 1067) on nymphal development time, fecundity and adult survival in Dichroplus maculipennis and Ronderosia bergi under laboratory conditions. It was observed that infection with 1×10³ conidia/ml altered nymphal development time, fecundity and adult survival in both species. Mortality of D. maculipennis during third through the last instar (sixth) was significantly higher among treated nymphs (66±3.8%) than in controls (15±1.7%). Similarly, mortality in R. bergi during third through the last instar (fifth) was higher in treated nymphs (71±2.8%) than in controls (19±1.5%). Nymphal development times of both infected D. maculipennis and R. bergi were longer than controls. On the other hand, among survivors of both the species, control adults lived longer than infected adults. Finally, control grasshoppers of both species were much more successful reproductively than infected grasshoppers.     

DNA synthesis in the neo-X neo-Y sex determination system of Dichroplus bergi (Orthoptera: Acrididae)

DNA replication in the neo-X neo-Y sex determining system was studied by means of tritiated thymidine and autoradiography. Asynchronous replication was found in the X arm of the neo-X and the long arm of the neo-Y. In addition, striking asynchrony was also found for short isopycnotic homologous regions at the distal end of the autosmal arm of neo-X and the short arm of neo-Y to which pairing during meiosis is restricted. These short regions are asynchronous with respect to the heterochromatic segments as well as to the remaining proximal region of the autosomal euchromatic arm of neo-X. This difference in replication pattern within the same chromosome arm may be related to a differentiation between regions which are homozygous in both sexes and regions which are hemizygous in males.This work was supported by U.S. Atomic Energy Commission Contract N AT (30-1) 3517 to Prof. F. A. Saez.     

The distribution of synaptonemal complex material in metaphase I bivalents of Locusta and Chloealtis (Orthoptera: Acrididae)

At metaphase I synaptonemal complex (SC) material is located in a continuous but irregularly shaped bundle between sister chromatids. Only at the site of a chiasma is it present between homologous chromosomes. When the chromosomes pull apart at anaphase I the SC material becomes rearranged into poly-SCs which dissociate from the chromosomes. The observations agree with previous reports that modified SCs may function in meiotic chromosome disjunction.