Spatio-temporal dynamics of a neutralized B chromosome in the grasshopper Eyprepocnemis plorans

Spatial and temporal patterns of frequency variation for a neutralized B chromosome in the grasshopper Eyprepocnemis plorans were analyzed along six transects in the east of Spain to explore possible factors affecting the population dynamics of this polymorphism. Three parameters were employed to quantify B frequency: prevalence, load and mean frequency. Of them, load seemed to be the less sensitive parameter, probably due to its small range of variation. Prevalence, however, shows ample variation, but the mean frequency of B chromosomes per individual is the best parameter to characterize B frequency. Only river transects revealed significant differences among populations, and the use of two geographic explicit approaches (Mantel test and distograms) revealed significant isolation by distance (IBD), especially at the Segura River mouth, presumably due to low gene flow and drift. No temporal trend was found in the Segura River transects, which is consistent with the slow changes in B frequency expected during the random walk for neutralized B chromosomes. But these transects showed a clear spatial pattern, with B1 showing lower frequency in the upper course of this river. The present results provide the first empirical evidence of IBD in the evolution of a neutralized B chromosome, and support the notion that B dynamics at this evolutionary stage is best explained by a metapopulation approach.     

Population dynamics of a selfish B chromosome neutralized by the standard genome in the grasshopper Eyprepocnemis plorans

Effects of the B chromosome polymorphism of the grasshopper Eyprepocnemis plorans were analyzed in two natural populations. Postmating sexual selection, female fertility, and survival were studied. The B chromosome lacks drive and has no detectable effects on fitness. A neutral B cannot invade a population and establish a polymorphism, but the confidence limits on our estimates cannot exclude the possibility that the polymorphism is maintained by a balance between weak drive and weak selection against individuals with two and three B's. However, other lines of evidence favor the following model of the dynamics of the B in E. plorans. In a newly invaded population, the B has substantial drive, but the evolution of drive suppressor genes in the A chromosomes neutralizes the B drive so that it becomes near-neutral and begins a random walk toward extinction by stochastic loss. Because the B is common by the time drive disappears, the random walk is likely to continue for a long time. If in the course of the random walk a variant B with greater drive appears, then it will displace the original variant, and a new cycle of drive suppression and drift to extinction occurs. A simulation model of this process suggested that the mean time to extinction is proportional to the two-thirds power of the population size; it is much less affected by subpopulation size or the number of populations in a subdivided population.     

The B chromosomes of the grasshopper Eyprepocnemis plorans and the intragenomic conflict

The grasshopper Eyprepocnemis plorans harbours an extremely widespread polymorphism for supernumerary (B) chromosomes, which is found in almost all circum-Mediterranean and Caucasian populations hitherto analysed. B chromosomes in this species have been shown to evolve through several stages of parasitic and near-neutral nature, presumably because of an arms race between the standard (A) and B chromosomes. This intragenomic conflict can either be solved with the extinction of the neutralised B chromosome or, more interestingly, with the replacement of the neutralised B by a mutant version being parasitic again and thus prolonging B chromosome life. This species thus provides a complete view of the long-term life-cycle of parasitic B chromosomes.     

The effect of B chromosomes on mating success of the grasshopper Eyprepocnemis plorans

The mating ability of E. plorans was tested in laboratory conditions in six experimental units composed of ten males and fifteen females during 31 days. When significant differences were found (three from the six cages, and in totals) they involved a decrease of matings involving males with B chromosomes. The same tendency seems to exist in females, but to a lesser extent, so that a significant effect is only detected when the totals are considered. Accessory chromosomes also delay, in both sexes, the occurrence of the first mating. No mating preferences depending on the number of Bs were detected.     

A single, recent origin of the accessory B chromosome of the grasshopper Eyprepocnemis plorans

B chromosomes are dispensable chromosomes found in >2000 eukaryotic species, usually behaving as genomic parasites. Most B chromosomes seem to be made up of the same kind of DNA sequences present in the A chromosomes. This sequence similarity makes it difficult to obtain specific molecular probes that may permit B-presence diagnosis without cytogenetic analysis. We have developed a sequence-characterized amplified region (SCAR) marker for B chromosomes in the grasshopper Eyprepocnemis plorans, which specifically amplifies a 1510-bp DNA fragment exclusively in B-carrying individuals. Fluorescent in situ hybridization and fiber FISH analyses showed that this marker is a tandemly repeated DNA sequence closely intermingled with 45S rDNA. PCR reactions showed the presence of SCAR-like sequences in the A chromosomes, but in two separate fragments, supporting the intraspecific origin of B chromosomes in this species. SCAR marker DNA sequence showed to be identical in B chromosome variants from several localities from Spain and Morocco, and it was very similar to those found in B chromosome variants from Greece and Armenia. This strongly suggests that this sequence was already present in the ancestral B chromosome of this species. In addition, the scarce sequence variation observed among several B variants from very distant populations suggests either a functional constraint or, more likely, a recent and unique origin for B chromosomes in this species.     

Parallel effects of a B chromosome and a mite that decrease female fitness in the grasshopper Eyprepocnemis plorans

The effects of a genomic parasite (a B chromosome) and an ectoparasite (a mite) on the fitness of the host (the grasshopper Eyprepocnemis plorans) have been analysed in 60 experimental females. These two parasites differ in their infectious transmission mode. B chromosomes are vertically transmitted from host-parents to offspring, but mites are horizontally transmitted from one grasshopper to another within the same generation. The transmission mode can influence the virulence of these parasites, so that it should be expected that B chromosomes would be less virulent than mites. However, as mite transmission is linked to host mobility, some attenuation is also expected. Four egg pods were analysed from each female, the first two egg pods were laid after a mating and the remaining two were not preceded by a mating. The results show that B chromosomes severely decrease the proportion of eggs containing an embryo (egg fertility), mainly from the second egg pod onwards. Mites also decrease egg fertility but, in addition, they produced a decrease in the rate of embryo production over time (embryo productivity), which might be derived from both the fertility decrease and a slight delay in egg production. The analysis of the relative effect of both parasites suggests that they have a synergistic effect on embryo clutch size and egg fertility. Possible mechanisms for the observed effects are discussed.     

Dynamics of sperm storage in the grasshopper Eyprepocnemis plorans

Abstract. Sperm storage duration in the grasshopper Eyprepocnemis plorans Charpentier (Orthoptera, Acrididae) was found to be 58.5 days but with a wide between-female variation (from 26 to 113 days). One copulation was enough to fertilize two to eight egg-pods and to produce 130 eggs (28–313) and 119 embryos (28–290), on average. Sperm availability, however, decreased progressively with time, so that the majority of females spent a long final period of their lives without laying any pod. Finally, sperm storage duration was positively correlated with clutch size and total production of eggs and embryos.     

Undertransmission of a supernumerary chromosome segment through heterozygous females possessing B chromosomes in the grasshopper Eyprepocnemis plorans.

The transmission ratio (ks) for a supernumerary chromosome segment was studied in a total of 54 heterozygous females collected from two Spanish natural populations of the grasshopper Eyprepocnemis plorans. Our analysis clearly demonstrated that ks is negatively dependent on the number of B chromosomes in the female. The possible mechanisms by which B chromosomes may cause undertransmission of the supernumerary segment, and the implications of this phenomenon for the maintenance of this extra chromosome segment, are discussed.     

Evolution of a complex B-chromosome polymorphism in the grasshopper Eyprepocnemis plorans

B-chromosome systems in several Spanish natural populations of Eyprepocnemis plorans are reported. The geographical distributions of the fourteen types of B chromosomes, which were classified according to size, C-banding pattern, and meiotic behaviour, are described. The results indicate a common origin for most types and a possible independent origin for a few secondary B variants. The origin and mechanisms of evolution of this B chromosome polymorphism are discussed.     

The B chromosome polymorphism of the grasshopper Eyprepocnemis plorans in North Africa. IV. Transmission of rare B chromosome variants

In addition to the principal B chromosome (B(1)) in Moroccan populations of the grasshopper Eyprepocnemis plorans, nine B chromosome variants appeared at low frequency. The transmission of five of these rare B chromosome variants through females was analysed in three natural populations. Sixteen controlled crosses provided useful information on the transmission of B(M2), B(M6) and B(M7) in Smir, B(M3) and B(M6) in SO.DE.A. (Société de Développement Agricole lands near Ksar-el-Kebir city), and B(M2) and B(M10) in Mechra, all located in Morocco. Since six female parents carried two different B variants, a total of 22 progeny analyses could be studied. Intraindividual variation in B transmission rate (k(B)) was observed among the successive egg pods in 26.7 % of the females, but this variation did not show a consistent temporal pattern. Only the B(M2) and B(M6) variants in Smir showed net drive, although variation was high among crosses, especially for B(M2). These two variants are thus good candidates for future regenerations (the replacement of a neutralized B, B(1) in this case, by a new driving variant, B(M2) or B(M6)) in Smir, the northern population where the B polymorphism is presumably older. The analysis of all crosses performed in the three populations, including those reported previously for the analysis of B(1) transmission, showed that the largest variance in k(B) among crosses stands at the individual level, and not at population or type of B levels. The implications of these findings for the occurrence of possible regeneration processes in Moroccan populations are discussed.     

Differences in ribosomal DNA distribution on A and B chromosomes between eastern and western populations of the grasshopper Eyprepocnemis plorans plorans

Distribution of ribosomal DNA (rDNA) on standard (A) and supernumerary (B) chromosomes of the grasshopper Eyprepocnemis plorans was analysed in specimens collected in Turkey and Armenia, belonging to the E. p. plorans subspecies, and in South Africa, belonging to the E. p. meridionalis subspecies. The latter individuals showed rDNA loci in chromosomes 9 and 11 only, whereas those from Armenia carried it in chromosomes 9 and 11 or else in chromosomes 9-11, depending on the population. The specimens from Turkey carried it in chromosomes 1, 9-11 and X. A comparison of this pattern with those previously observed in populations from Spain, Morocco, and Greece (belonging to E. p. plorans) suggests the existence of two evolutionary patterns in rDNA chromosome location in A chromosomes of this subspecies: eastern populations showing rDNA restricted to the small (9-11) chromosomes (as in E. p. meridionalis and other closely related taxa within the Eyprepocneminae subfamily) and western populations carrying rDNA in most A chromosomes (Spain) or all of them (Morocco). The intermediate pattern discerned in geographically intermediate populations (in Greece and Turkey), with rDNA also being located on the X chromosome, suggests a possible east-west cline. Additional support for east-west differentiation in the rDNA location pattern comes from the analysis of B chromosomes. In eastern populations, including Daghestan, Armenia, Turkey, and Greece, B chromosomes are composed mostly of rDNA, whereas in western populations (Spain and Morocco) they contain roughly similar amounts of rDNA and a 180-bp tandem repeat (satDNA), the latter being scarce in eastern Bs.     

The B chromosome polymorphism of the grasshopper Eyprepocnemis plorans in North Africa: III. mutation rate of B chromosomes

B chromosome variation in nine Moroccan populations of the grasshopper Eyprepocnemis plorans was analysed for 3 consecutive years. In addition to B1, which was the predominant B chromosome in all nine populations, we found 15 other B variants, albeit at very low frequency. Eight variants were found in adults caught in the wild, four appeared in adults reared in the laboratory and seven were found in embryo progeny of controlled crosses between a 0B male and a B-carrying female. Some variants were found in more than one kind of material. At least the seven B variants that appeared in embryo progeny of females carrying a different B type arose de novo through mutation of the maternal B chromosome. The mutation rate of B chromosomes was 0.73%, on average, which explains the high variety of morphs and banding patterns found. The most frequent de novo mutations observed in these chromosomes were centromere misdivision with or without chromatid nondisjunction, which generates iso-B-chromosomes or telocentric Bs, respectively, as well as translocations with A and B chromosomes and deletions. But the whole variation observed, including that found in adult individuals, suggests that other mutations such as duplications, inversions and centric fusions do usually affect B chromosomes. Finally, B chromosome mutation rate was remarkably similar in both Moroccan and Spanish populations, which suggests that it might be dependent on B chromosome intrinsic factors.     

Gene expression changes elicited by a parasitic B chromosome in the grasshopper Eyprepocnemis plorans are consistent with its phenotypic effects

Chromosoma - Parasitism evokes adaptive physiological changes in the host, many of which take place through gene expression changes. This response can be more or less local, depending on the organ...     

The b chromosome polymorphism in Armenian and Turkey populations of the grasshopper Eyprepocnemis plorans plorans (Charpentier) (Orthoptera, Acrididae, Eyprepocnemidinae)

Karyotype analysis of the grasshopper Eyprepocnemis plorans samples derived from Armenian and Turkey populations was performed using C-banding technique. Chromosome polymorphism was associated only with B chromosomes that were revealed in all studied populations. Six new B chromosome morphotypes were described. Four morphotypes were found only in Armenian populations. One morphotype was revealed only in Turkey populations. One B chromosome morphotype was present in Armenian and Turkey populations. B chromosomes derived from Asian populations consisted mostly from C-positive regions. In some of the B chromosomes small C-negative regions were also registered. Morphotypes of the B chromosomes derived from Armenian and Turkey populations drastically differed from the B chromosomes described in the Iberian Peninsula and North African populations. In contrast to the B chromosomes from Armenian and Turkey populations the B chromosomes from Spain populations contained C-positive and C-negative regions alternated in their arms.     

Spontaneous translocations between B chromosomes and the normal complement in the grasshopper Eyprepocnemis plorans

A translocation between a B chromosome and a medium-size autosome and a centric fusion between the B and the X chromosome were found in two different natural populations of Eyprepocnemis plorans. The cytological behaviour of both interchanges is described and discussed. These interchanges are highly asymmetrical and may be the beginning of an integration of B chromosomes into the genome of this species.     

Meiotic drive against an autosomal supernumerary segment promoted by the presence of a B chromosome in females of the grasshopper Eyprepocnemis plorans

Twenty-seven out of 50 progeny analyses performed with specimens of the grasshopper Eyprepocnemis plorans were informative about the transmission of a supernumerary heterochromatic chromosome segment. The simultaneous presence of a B chromosome in some of the parents involved in the crosses permitted us to test the relationship between both types of supernumerary heterochromatin with respect to their transmission. The results demonstrated that the supernumerary segment is partly eliminated through females possessing B chromosomes. The implications of this in relation to the occurrence of the extra segment in natural populations are discussed.by S.A. Gerbi     

Detection of B chromosomes in interphase hemolymph nuclei from living specimens of the grasshopper Eyprepocnemis plorans

The two most important evolutionary properties of B chromosomes are their transmission rate (which suggests their selfishness when significantly higher than 0.5) and their net effects on carrier fitness (usually negative for parasitic Bs). The study of transmission rate unavoidably requires the analysis of many controlled crosses in order to accurately measure population average transmission rate. Therefore, getting a marker closely associated to B presence is of crucial importance to alleviate the load of performing many useless crosses between lacking B individuals. After investigating several cytogenetic techniques on several tissues that may be sampled without drastically damaging live specimens of the grasshopper Eyprepocnemis plorans, we report here the excellent results provided by the CMA3 fluorescence and C-banding techniques applied to hemocyte nuclei. These cells may be easily obtained from both males and females and provide information on B presence even during the interphase stage. The two cytogenetic techniques take advantage of the heterochromatic nature of the B chromosomes, so that Bs made predominantly of ribosomal DNA are revealed by CMA3 as bright bodies in the interphase hemocytes, and Bs mostly made of satellite DNA are visualized by C-banding as intensely stained bodies in these cells.     

Physical mapping of rDNA and satDNA in A and B chromosomes of the grasshopper Eyprepocnemis plorans from a Greek population

Adult males and females of the grasshopper Eyprepocnemis plorans from a Greek population were analysed by C-banding, silver impregnation and double FISH for two DNA probes, i.e. ribosomal DNA (rDNA) and a 180-bp tandem repeat DNA (satDNA). This population shows characteristics of rDNA location in A chromosomes that are intermediate between those previously reported for eastern (Caucasus) and western (Spain and Morocco) populations. The four rDNA clusters revealed by FISH in chromosomes X, 9, 10 and 11 in Greek specimens imply two more than the two observed in chromosomes 9 and 11 in the Caucasus, but less than the 12 observed in all chromosomes in Morocco. Remarkably, the X chromosome bears one of the new rDNA locations in Greece with respect to the Caucasus, but it appears to be inactive, in contrast to X chromosomes in western populations, which are usually active. B chromosomes were very frequent in the Greek population, and three variants differing in size were observed, all of these being largely composed of rDNA, with the exception of a small pericentromeric satDNA cluster. The high B frequency suggests that B chromosomes in this population might behave parasitically, in resemblance to Bs in western populations.     

Changes in NOR activity pattern in the presence of supernumerary heterochromatin in the grasshopper Eyprepocnemis plorans

Nucleolus organizer region (NOR) activity was analysed in four types of males of the grasshopper Eyprepocnemis plorans, possessing two kinds of supernumerary heterochromatin: a B chromosome and a supernumerary chromosome segment proximally located on the smallest autosome (S11). In males lacking extra heterochromatin, the four active NORs located on the S9, S10, S11, and X chromosomes showed independent activity patterns, but several kinds of dependence appeared in the presence of supernumerary heterochromatin. Furthermore, temporal changes in NOR activity were observed during the first 2 weeks of adult life in standard males but not in males carrying supernumerary heterochromatin. It is suggested that all these effects are related to the DNA content of both types of extra heterochromatin.