Maintenance of a “Parasitic” B Chromosome in the Grasshopper Melanoplus Femur-Rubrum

MAINTENANCE OF A “PARASITIC” B CHROMOSOME IN THE GRASSHOPPER MELANOPLUS FEMUR-RUBRUM

UZI NUR Departmant of Biology, Uniuersity of Rochester, Rochester, New York 14627 Manuscript received April 20, 1977 Revised copy received August 1, 1977

ABSTRACT

About 10-15% of the males and females of the grasshopper Melanoplus femur-rubrum collected near Rochester, New York, possessed a supernumerary B chromosome. The frequency of the B chromosome remained fairly constant during the years 1971-1974. The B chromosome was shown previously to be transmitted at a rate of about 0.5 and 0.8 by 1B males and females, respectively. This study was designed to determine the forces preventing the B chromosome from increasing in frequency due to the high rate of transmission by the females. Eighty inseminated females collected in the wild were analyzed cytologically together with their embryos (10-20 per female). Ten of the 80 females had a B chromosome, and they transmitted it at a rate of about 0.75. Among the 983 embryos analyzed, 0.141 had one B, 0.007 had two, and the mean number of B chromosomes per embryo was 0.155. The frequency of the B chromosome in the sperm pool (0.061) was consistent with a 0.5 rate of transmission. Individuals with two B chromosomes apparently have low viability, because about six were expected, but none was found among 851 adult males and females examined. The data suggest that the viability of the 1B individuals was only about 0.86 that of the OB individuals. There was no evidence that the B chromosome increased the fecundity of either the 1B males or females. It was concluded, therefore, that the B chromosome reduced the fitness of all the individuals carrying it and was thus “parasitic,” and that it was maintained in the population only because of its high transmission rate. The maintenance of other B chromosomes with high transmission rates is reviewed.

chromosomes (supernumerary chromosomes) are present in some of the indi- Bviduals of a population, but not in others, are not homologous to the regular chromosomes, and usually have little effect on the phenotype. B chromosomes are present in many animal and plant species (WHITE1973; JONES1975). Some of the B chromosomes are mitotically unstable (NUR1963,1969a; WHITE1973) or undergo nondisjunction during meiosis (JONES1975), and in some species these unusual types of behavior may serve as accumulation mechanisms that, like meiotic drive, would tend to cause the frequency of the B chromosomes to increase from generation to generation. In several species that possessed an accumulation mechanism, the frequency of the B chromosomes remained stable over several generations or years (NUR196910; HEWITT1973b), and this stability could result only from selection against some of the individuals with B chromo-

Genetics 87: 499-512 November. 1977. 500 U. NUR somes. The selection against them may take place in two ways, which were termed by WHITE(1973) the “heterotic” and “parasitic” models. According to the former model, the fitness of individuals with one B chromosome (1B individuals) is higher while that of those with higher numbers of B chromosomes is lower than OB individuals. According to the “parasitic” model, the mean fitness of all the classes of individuals with B chromosomes is lower than that of OB individuals. More complex models in which, for example, the B chromosomes are “heterotic” in one sex but “parasitic” in the other are possible. It is also possible that different models may apply to the same B chromosome in different populations, to B chromosomes of different species, or to different B chromosomes of the same species. It has been pointed out earlier (NUR1969a; Lucov and NUR 1973) that in species like the grasshopper Melanoplus femur-rubrum in which the frequency of B chromosomes is fairly low (0.10-0.15) and the tendency to accumulate is fairly high (the combined rates of transmission in both sexes is 1.2-1.3), the B chromosome cannot be “heterotic” in both sexes, because even if 2B individuals were effectively lethal, their death would not remove enough B chromosomes to offset the tendency to accumulate. The possibility could not be ruled out, however, that in such species the B chromosome was “heterotic” in one sex but “parasitic” in the other. The aim of this study was, thus, to try to obtain information about the fitness of OB, 1B and 2B males and females of Melanoplus femur- rubrunz in order to determine whether in this species the B chromosome is maintained according to the “heterotic,” the “parasitic,” or a more complex model.

MATERIALS AND METHODS

The males and females analyzed were collected in a field and an adjacent lawn near the Whipple Park housing project of the University of Rochester, Rochester, New York. The 1971, 1972, and 1973 collections were made from the same area, which was about 30 x 120m. In 1974, part of the field was plowed and this part and the adjacent lawn had very few grasshoppers. Thus, the collection was restricted to an area of about 30 x 30 m, which comprised the southwest section of the area from which previous collections were made. In 1973, adult males were collected during September and the first week of October. Sixty- eight adult females were collected on September 12, 1973, and placed individually in small cardboard cages with lettuce, oat bran and damp sand. In the next six days six females died and 32 laid one egg pod each. Because of the possibility that additional mortality might invali- date the sample, all the females were killed and fixed. A second group of 79 females was collected on September 25, 1973, and maintained as before. Only three females died during the next 16 days, at which time 51 of the females had oviposited, and these were fixed for analysis as soon as laying was detected. The nonlaying females were all fixed on the 16th day after their collection. Most of the eggs of two of the 83 laying females did not contain embryos. The eggs of one female were assumed to have developed parthenogenetically because all the embryos were very small, relative to those which were laid on the same day by other females, and some of the embryos contained haploid cells. Thus, the eggs of those three females were not included in the analysis. The eggs of the remaining 80 females were allowed to develop for 10-14 days and the embryos were then dissected out, treated with a hypotonic solution containing colchicine (Lucov and NUR1973) and then fixed. The laying females were examined cytologically prior to the dissection and the fixation of their embryos. When a female was four,d to possess a B chromosome, about 20 of her embryos “PARASITIC” B CHROMOSOME 501 were treated and fixed. For the OB females, either 10 or about 20 embryos were treated and fixed, depending on the time available that day. In 1974, immature and adult males and immature females were collected between August 23 and September 16. The adult males were fixed immediately. The immature males and females were kept in the laboratory until they became adults, during which time a few died. The rest were then fixed and analyzed cytologically.

RESULTS The apperrrance and behavior of the B chromosome The B chromosome was about the size of several of the medium-sized regular chromosomes. Unlike all the regular chromosomes, however, it was metacentric, and as such could be readily identified during mitosis (Figures 1 and 2). In 1B males, during prophase I of meiosis, the B and the X chromosome both appeared positively heteropycnotic (heterochromatic), but the B was slightly shorter than the X chromosome (Figure 3). The B chromosome moved to one

FIGURE1.-Mitosis in a female embryo with one metacentric B chromosome (lower center) and 24 regular chromosomes. FIGURE2.--Mitosis in a male embryo with two metacentric B chromosomes and 23 regular chromosomes. FIGURE3.-Diplotene-diakinesis in a spermatocyte with one heterochromatic B chromosome (lower right). The heterochromatic X chromosome is at the lower left.

FIGURE4.-Diplotene-diakinesis in a spermatocyte with two small heterochromatic BT chromosomes. One BT chromosome is near the X chromosome and the other is slightly above it. 502 U. NUR of the poles in anaphase I and divided regularly during anaphase 11. The B chromosome was mitotically stable, and there was no evidence to suggest that it tended to be lost at any time between the zygote and the spermatids. As has been reported by Lucov and NUR(1973) , during anaphase I of oogenesis the B chromosome tended to move preferentially into the secondary oocyte, rather than into polar body I, and this tendency apparently caused it to be transmitted at a rate which was significantly higher than the expected rate of 0.5. The frequency of the B chromosome among adults The results of the €our years of sampling the Whipple Park population are presented in Table 1. The frequency of the B chromosome among both sexes ranged from 0.107 to 0.136. In three of the four years, its frequency among females was lower than among males, but none of the differences was statistically significant. The frequency of the B chromosome in 1971 (0.107) was somewhat lower than in the subsequent years, but at least for the years 1972- 1974 its frequency remained fairly constant. One unexpected result of the sampling was the increase in the frequency of a small extra chromosome from 1971-1973 to 1974. In its behavior in males, this chromosome appeared to be similar to a small B chromosome described previously by STEPHENSand BREGMAN(1972) and designated by them BT. Like BT, the extra chromosome was heterochromatic in prophase I of spermatogenesis (Figure 4) and was mitotically unstable. During mitosis, the extra chromosome closely resembled one of the six small autosomes. The large difference between the frequency of the BT chromosome in 1974 (about 7%) and in 1973 (not observed in the males, present in 2% of the females and in about 3% of the embryos) may have been the result of the fact that in 1974 the males and females were collected from only part of the area from which they were collected previously (see MATERIALS AND METHODS). The higher fre-

TABLE 1 The frequencies of adult males and females with and without a metacentric B chromosome in the Whipple Park population

Males Females Both sexes Year N OB 1B N OB 1B N OB 1B 1. 1971 143 0.867 0.133 156 0.917 0.083 299 0.893 0.107 2. 1972 51 0.843 0.157 42 0.881 0.119 93* 0.860 0.140 3. 1973 134 0.873 0.127 138 0.855 0.143 2721 0.864 0.136 4. 1974 145 0.848 0.152 42 0.929 0.071 187% 0.866 0.134 Totals (1-3) 328 0.866 0.134 336 0.887 0.113 664 0.877 0.123 Means (1-3) - 0.861 0.139 - 0.884 0.116 - (1.872 0.128

The data for 1971 and 1972 are from Lucov and NUR(1973). The reasons for not including the 1374 data in the totals and means are discussed in the text. * Including one female with one telocentric BT. Including three OB females with BT chromosomes. $Including eight 01B males, three OB females, one 1B male and one 1B female with BT chromosomes. “PARASITIC” B CHROMOSOME 503 quency of the BT chromosome in the 1974 sample suggested that the animals sampled in 1974 were a distinct subpopulation, and for this reason in Table 1 the 1974 data were not combined with those of the other years. The frequency of the B chromosome among the embryos The frequency of the B chromosome among the embryos of the 80 laying females is given in Table 2 and summarized in Table 3. Among the embryos of all but one of the 1B females, the frequency of embryos with B chromosomes was 0.7 or more, and among those of the OB females, it was 0.55 or lower. Each of the females laid one egg pod, and in almost all the pods all the eggs contained embryos. Thus, differential mortality of embryos of different karyotypes could not have had a major effect on the observed frequency of the B chromosome among the embryos. TABLE 2 The frequencies of the uarious karyotypes among 80 females which were inyeminateed in the wild and their embryos

Female’s Embryos/ Frequency of embryos B’s per By’s per Female karyokype efific OB 1B 2B embryo embryo N 1 1B 26/27 0.087 0.478 0.435* 1.348 23 2 1B 39/39 0.100 0.900 0.900 20 3 1B 14/36 0.214 0.714 0.857 14 4 1B 32/33 0.150 0.850 0.850 20 5 1B 37/37 0.150 0.850 0.850 20 6 1B 31/31 0.200 0.800 0.800 20 7 1B 36/36 0.250 0.750 0.750 20 8 1B 34/34 0.250 0.750 0.750 20 9 1B 3 7/41 0.300 0.700 0.700 20 10 1B 32/34. 0.762 0.190 0.286 21 11 OB 33/33 0.450 0.550 0.550 20 12 OB 17/19 0.600 0.400 0.400 10 13 OB 27/23 0.600 0.400 0.400 10 14 1BT 26/31 0.611 0.389 0.389 18 15 OB 35/35 0.632 0.368 0.368 19 16 O’B 18/18 0.700 0.300 0.300 10 17 OB 28/28 0.700 0.300 0.300 10 18 OB 32/32 0.700 0.300 0.300 20 19 OB 21 /2 1 0.762 0.238 0.238 21 20 OB 34/35 0.800 0.200 0.200 20 21 OB 28/29 0.800 0.200 0.200 20 22 OB 34/34 0.850 0.150 0.150 20 23 OB 401/40 0.900 0.100 0.100 20 24 OB 32/33 1.000 - - 10 25 1BT 34/34 1.000 - - 10 26-80 OB l.m - - 547 Means and sum 0.857t 0.136 0.150 983

Embryos/eggs represents the number of eggs with embryos among the eggs in the egg pod. N is the number of embryos analyzed cytologically. The data from 55 females (26-80) whose embryos lacked either B or BT chromosomes were combined. * Including one embryo with 36 + 3B chromosomes. t Including one embryo with 35 and three with 36 chromosomes. 504 U. NUR

In this species male are XO, 212 = 23, and females are XX, 2n = 24. The X is one of the medium-sized chromosomes (Figures 3 and 4). Among the 198 embryos of the ten 1B females listed in Table 2, 52% had 17 medium to large acrocentric chromosomes and these embryos were considered to be males; the rest had 18 such chromosomes and they were considered to be females. Among the male embryos, 75% had B chromosomes and among the female embryos 75.6% had B chromosomes. Among the 218 embryos of the 13 OB females with at least one embryo with a B chromosome (Table 2, Females 11-23), 51 % were males and the frequencies of embryos with B chromosomes among the male and female embryos were again about equal-28% and 32% respectively. Two of the laying females and about 3% of the embryos possessed seven small chromosomes (instead of six), and were thus classified as possessing the BT chromosome (Table 2). The frequency of the B chromosome in the sperm pool The frequency of the B chromosome in the sperm pool of the population can be estimated from the frequency of embryos with B chromosomes among the offspring of the OB females. On the basis of the data from the first 10 embryos analyzed from each of 67 OB females and the 9 embryos analyzed from three other OB females, 6.1% of the sperm carried a B chromosome (Table 3, second row). For some of the OB females, data were available from more than 10 embryos, but these data were not used in order not to bias the analysis, because the decision to analyze 10 additional embryos from some of these females was based on the finding of at least one 1B embryo among their offspring, and thus these additional embryos did not represent a random sample. In controlled crosses five 1B males were found to transmit their B chromosome at a rate of about 0.5 (Lucov and NUR 1973). The findings that 12.7% of the 134 males examined in 1973 (Table 1) and 6.1 % of the sperm carried a B chromosome are thus consistent with the results of the controlled crosses, since they suggest a rate of transmission of about 0.48. Thirteen of the OB females had at least one embryo with a B chromosome, and these females must have mated with B-possessing males. The average frequency of the B chromosome among

TABLE 3 Summary of the analysis of the 80 laying females and their embryos

~~ Frequency of karyotypes Females OB 1B 2B N Mean B Embryos of 1B females 10 0.247 0.698 0.055 198 0.808 Embryos of OB females 70 0.939 0.061 0.0 697 0.061 Embryos of all females 80 0.852 0.141 0.007 895 0.155 Laying females 80 0.875 0.125 0.0 - 0.125 Survival-zygotes to adult females - 1.000 01.863 0.0 - -

The data about the embryos of the OB females are based only on the first 10 embryos analyzed from each of these females. The calculated survival from zygotes to adult females is based on the assumption that the females developed from zygotes similar to those giving rise to the 895 embryos. N-number of embryos analyzed. “PARASITIC” B CHROMOSOME 505 the first 10 embryos analyzed from these 13 females, however, was only 0.328 (rather than 0.5). This low value was almost certainly the result of multiple matings involving both OB and 1B males (rather than a low transmission rate), since multiple matings were often observed in the lab. Female transmission and eggs per pod The average rate at which the B chromosomes were transmitted by the females, kf,may be estimated by subtracting the frequency of the B chromosomes in the sperm pool (0.061) from the frequency of the B chromosomes among the embryos of the 1B iemales (0.808, Table 3). Such a calculation gives a rate of transmission of 0.747. This rate is significantly higher than 0.5, but is not significantly different frcm the rate of 0.817 4 0.052 obtained in controlled crosses (Lucov and NUR1973). The 10 1B females laid an average of 34.8 eggs per pod, while 72 OB females laid 30.7 eggs per pod. The differences is not statistically significant (t= 1.808; p > 0.05). The frequency of 2B individuals Among the 851 adults collected at Whipple Park and analyzed cytologically (Table l), none had two B chromosomes. Among the 983 embryos analyzed, however, 0.007 had two. This value is approximately that to be expected in a population in which 0.125 of the males and females had one B chromosome and in which the males transmitted it at a rate of about 0.5 and the females at a rate of about 0.75 (0.125 x 0.5 x 0.125 X 0.75 = 0.006). When the expected frequency of 2B individuals is 0.007, the probability of not finding 2B individuals among 851 individuals is (0.993)s51= 0.006. When the probability of survival of the 2B individuals is 0.5 (relative to that of the other individuals), the probability of not finding any 2B individuals is (0.9965)s51= 0.05. Thus, the absence of 2B adults suggests that the viability of 2B individuals is less than 0.5.

DISCUSSION The maintenance of the B chromosome in NI. femur-rubrum The results presented in the previous sections suggested the following conclusions: (1) The frequency of the B chromosome in the population was stable. (2) The frequencies of the B chromosome were approximately the same in the males and females among the embryos and also among the adults. (3) The 2B individuals had low survival. (4) The number of eggs in the pods laid by OB and 1B females was about the same. (5) The frequency of the B chromosome among the sperm (0.06) was approximately that expected from the observed frequency of 1B males (0.127) and their previously determined rate of transmission (about 0.5). Thus, the B chromosome apparently had little or no effect on male fertility and fecundity. (6) The 1B females transmitted the B chromosome at a rate of about 0.75. (7) The frequency of the B chromosome was about 0.125 among adult males and females and about 0.155 among the zygotes. Thus, the B increased in frequency from the adults to the zygotes of the next generation by about 0.03. (8) The selection against the 2B individuals could not 506 U. NUR have removed more than about 0.007 x 2 = 0.014, or only about 50% of the increase in the frequency of the B chromosome due to the high transmission by the 1B female. Thus, the rest of the elimination had to be through the reduction in the fitness of at least some of the 1B individuals. On the basis of the frequency of the 1B individuals among the females (0.125) and that among the embryos (0.141) , it was calculated (Table 3) that the survival of the 1B females was only about 0.86 that of the OB females. Among both the embryos and the adults, the frequency of the 1B males was about the same as that of the 1B females, and thus the relative survival of the 1B males had to be also about 0.86. But since there was no evidence that the B chromosome increased the fertility or the fecundity of either the 1B males or the 1B females, relative to that of the OB individuals, it must be concluded that the B chromosome is of the “parasitic” type. The same conclusion was reached previously by LUCOV and NUR (1973) on the basis of the low frequency of the B chromosome in the population and its rate of transmission by the two sexes. At this point it may be noted that although the frequency of the 1B embryos among the non-2B embryos (0.142) is not significantly greater than that among either the adult males or the adult females (about 0.125), the increase is almost certainly real for the following reasons. The contribution of the 1B males to the non-2B embryos (0.061 B chromosomes per embryo) was very close to that expected (0.0635) on the basis of the frequency of these males (0.127) and their observed rate of transmission (0.5). On the other hand, the contribution of the 1B females to the non-2B embryos (0.081) was clearly greater than that to be expected (0.0625), since the frequency of the 1B embryos among the non- 2B embryos of the ten 1B females (0.742 i. .063) was significantly greater than the expected value of 0.5. It may be concluded, therefore, that the increase in the frequency of the 1B embryos was real, that the stability in the frequency of the B chromosome was at least in part due to selection against 1B males and females, and that this selection was mostly through the reduction in the viability of the 1B karyotype. The maintenance of other B chromosomes Whether in a given population the B chromosome is “heterotic” or “parasitic” is likely to depend on whether the average rate of its transmission in both sexes is less than 0.5, 0.5, or higher than 0.5. It is likely that a B chromosome with an average rate of transmission of less than 0.5 would confer an advantage on at least some of the individuals carrying it, or else it would be quickly eliminated. When the average rate is exactly 0.5, it is conceivable that such a B chromosome would have no effect on fitness. It is more likely, however, that for the B chromosome to be maintained over many generations, some frequency-dependent selection must be involved in which it would confer a selective advantage on one (or more) of the karyotypes and disadvantage on the others. Only when the average rate of transmission of the B chromosome in both sexes is greater than 0.5, as by an accumulation mechanism or meiotic drive, might it be possible for the B chromosome tQreduce the fitness of all the individuals carrying it, thus being “PARASITIC” B CHROMOSOME 507 “parasitic,” and still be maintained. A B chromosome with an accumulation mechanism, however, might also be maintained if it increases the fitness of some of the individuals carrying it, relative to that of OB individuals, and thus be “heterotic,” as long as selection against the remaining individuals with B chromosomes is sufficiently strong to offset the increase in the frequency of the B chromosome through both the accumulation mechanism and the superiority of some of the B-carrying individuals. Thus, it might be of interest to determine whether in other species with B chromosomes that are known to have an accumulation mechanism the B chromo- wmes are LLpara~iti~7’or “heterotic.” In order to demonstrate, however, that a particular B chromosome is “heterotic,” it is not enough to show (see review in JONES1975) that it affected some of the individuals carrying it in a way which might be considered adaptive. For example, in Lolium prenne, the B chromosome increases the survival of some of the plants carrying it under certain experimental conditions (REES and HUTCHINSON1973; HUTCHINSON1975). In- stead, it is necessary to demonstrate that the B chromosome increases the fitness of at least some of the individuals carrying it under natural conditions. Unfor- tunately, there are only a few cases in which this has been attempted and these will now be reviewed. The first analysis of this kind was that carried out by KIMURAand KAYANO (1961) on a B chromosome (termed f) in Lilium callosum. The authors used information about its frequency in several natural populations, its rate of transmission through the pollen and the egg, and its effect on pollen and seed fertility under experimental conditions to calculate the expected viabilities of the various karyotypes, and by combining the information about the viability and €ertility concluded that “whatever modification is made in the selective pattern by local ecological conditions, the conclusion is inevitable that in Lilium callosum the f chromosome is predominantly deleterious and it may only be kept in the population by preferential segregation in embryo sac mother cells.” On the basis of KIMURAand KAYANO’Sdata, the relative fitness of the OB, 1B and 2B karyotypes may be calculated to be 1.00, 0.94, and 0.39, respectively. Thus, the B chromosome of this species appears to be “parasitic.” The second B chromosome for which such an analysis was carried out was that of the mealybug Pseudococcus obscurus (NUR 1966a). The analysis was similar to that of the present study in that females inseminated in the wild were analyzed cytologically together with their embryos, and the rates of transmission of the B chromosomes by both sexes were determined in controlled crosses. The analysis indicated that the B chromosome had little or no effect on the females, but reduced the average fitness of all the males carrying it. This conclusion was then confirmed by allowing OB females to mate with wild males and analyzing their offspring (NUR 196913). In the latter study, the relative fitness of males with 0, 1, 2, 3. and 4 or more B chromosomes was determined to be 1.00, 0.64, 0.56, 0.38, and 0.2, respectively (NUR 1969b). Under laboratory conditions it was also established that a single B chromosome reduced the rate of development 508 U. NUR of the males and the number of sperm produced (NUR 1966b). Thus, this B chromosome is clearly “parasitic.” The grasshopper Myrmeleotettix maculatus The only other B chromosome with an accumulation mechanism for which an attempt was made to determine the fitness of the various karotypes, and thus the nature of the B chroinosome, is that of the grasshopper M. maculatus (HEWITT 1973a,b, 1976; ROBINSONand HEWITT1976). HEWITT(1973a,b) determined the rate of transmission of the B chromosome in a series of crosses involving individuals from two populations from Great Britain, Talybont and Foxhole, and then used these rates and the observed frequencies of the various karyotypes among adult males to calculate the expected fitness of the various karyotypes relative to that of the OB type. For Talybont the rate of transmission of 1B males, 1B females and 2B males were reported to be 0.496, 0.902, and 0.442, respectively, and the fitnesses of the lB, 2B and 3B karyotypes relative to that of the OB type were calculated to be 0.81, 0.73 and 0.07, respectively. Thus, the B chromosome of this population tended to accumulate and was apparently of the “parasitic” type. The Foxhole population of M. MacuZatus is of special interest because on the basis of its analysis ROBINSONand HEWITT(1976) concluded that the B chromosome was not parasitic, and on the basis of HEWITT’S(197313) analysis of the effect of the B chromosome on survival, it might even be classified as “heterotic.” Further analysis of the data, however, suggest that it might still be “parasitic.” The rates of transmission of the 1B males, 1B females, 2B males and 2B females of the Foxhole population were reported by HEWITT(197313) to be 0.366, 0.567, 0.302 and 0.625, respectively, and thus they were significantly lower than those from Talybont. The analysis of the data available about the Foxhole population is presented in Table 4 in the form of four sets of comparisons. The first set is that given by HEWITT(1973b). The analysis suggests that the B chromosome is “heterotic” because the survival of the OB individuals is less than that of both the 1B and 2B individuals, and according to ROBINSONand HEWITT (1976) there is no clear evidence of differential fecundity between the three karyotypes. Thus, it follows that the fitness of the OB individuals is less than that of at least some of those with the B chromosome. The second comparison is beween the 267 embryos obtained from eggs collected in the autumn, and adult males. As may be seen from the second comparison, the frequency for the B chromosome in the autumn eggs (0.753) is much higher than that among the adults (0.601) and this difference is statistically significant (t= 2.5; p < 0.02). Since the frequency of the B chromosome in the population appears to be stable (ROBINSONand HEWITT1976), the adults presumably developed from embryos similar to those collected in the autumn, and thus it is possible to calculate the relative survival of the various karyotypes from embryos in the autumn to the adults. According to such a calculation (Table 4, second comparison) the survival of the lB, 2B, 3B individuals was 0.89, 0.71 and 0.0 relative to that of the OB individuals. Thus, the B chromosome “PARASITIC” B CHROMOSOME 509

TABLE 4 Calculated relatiue suruiual of the uarious karyotypes in the Foxhole population of the grasshopper Myrmeleotettix maculatus

Frequency of karyotypes N OB 1B 2B 3B+4B Mean B 1. Expected zygotes (HEWITT1973b) - 0.535 0.383 0.077 0.005 0.553 Adult males (HEWITT1373b) 281 0.495 0.400 0.096 0.0 0.601 Survival-zygotes to adults (HEWITT1373b) 1.000 1.169 1.418 0.0 0.601 2. Autumn eggs (ROBINSONand HEWITT1976) 267 0.408 0.457 0.112 0.023 0.753 Adult males (HEWITT1973b) 281 0.495 0.409 0.096 0.0 0.6011 Survival-autumn eggs to adults 1.000 0.890 0.710 0.0 - 3. Expected zygotes (HEWITT1973b) 0.535 0.383 0.0877 0.005 0.553 Autumn eggs (ROBINSONand HEWITT1976) 267 0.408 0.457 0.112 01.023 0.753 Survival-zygotes to autumn eggs - 1.0100 1.560 1.910 6.03 - 4. Expected zygotes (k, = 0.48; k, = 0.78) - 0.420 0.420 0.142 0.0118 0.758 Adult males (HEWITT1973b) 28 1 0.495 0.409 0.096 0.0 0.601 Survival-zygotes to adults 1.000 0.830 0.570 0.0

The expected zygotes are either those calculated by HEWITT(197313) on the basis of controlled crosses, or are those calculated on the assumption that the rates of transmission of the B chromosome by the males and females (k, and kr) are 0.48 and 0.78, respectively.

appears to reduce the survival of individuals carrying it between eggs in the autumn and the adults, and the reduction increases with the increases in the number of B chromosomes. This result is of special interest because it is based on two observed frequencies and not on a comparison involving expected zygotes. Before one draws conclusions about the effect of the B chromosome on survival, however, it is also necessary to know its effect on survival between the zygotes and the autumn eggs, and this was calculated in the third comparison of Table 4. It may be noted here that its frequency in the autumn eggs (0.753) is much higher than that among the expected zygotes. ROBINSONand HEWITT (1976) commented on this increase as follows: “This difference in the observed B frequency and that predicted could be due to nonrandom mating or differential fecundity of the different karyotypes, but there is no clear evidence for either of these from our investigations to date. . . . However, the most obvious possible cause is differential selection for B-containing embryos.” The authors did not calculate the intensity of selection required to bring about this increase, but this was calculated in the third comparison of Table 4. From such a calculation, it may be concluded that for differential selection to account for the observed frequencies of the various karyotypes among the autumn eggs, the OB embryos would have to survive very poorly relative to those with B chromosomes and’ that the survival would have to improve with an increase in the number of B chromosomes. This conclusion, however, presents a serious difficulty because it would require strong differential selection between egg laying and the sampling, 510 U. NUR and according to the data of ROBINSONand HEWITT(1976), among the autumn eggs the frequency of eggs without embryos, and thus mortality, was quite low. One way to explain the observed increase in the frequency of the B chromosome among the autumn eggs relative to that among the adult males is to assume that its transmission was considerably higher than that determined by HEWITT in controlled crosses. This assumption is strengthened by HEWITT’S(1976) report that the transmission of the B chromosome by females from another population from East Anglia, Lakenheath Warren (which according to Figure 1 of ROBINSONand HEWITT(1976) is only about 3 km from Foxhole), is considerably higher than by those from Foxhole. Thus, according to HEWITT(1973b), the rate of transmission by the 1B females from Foxhole was about 0.6, while in the females from Lakenheath Warren it was 0.78 (HEWITT1976). In order to explore this alternative further, the expected frequencies of the various karyotypes among the zygotes were calculated under the assumption that the transmission of the B chromosome by the females from Foxhole was also 0.78. In the males the rate of transmission was assumed to be 0.48. This value of 0.48 is still lower than that reported by HEWITTfor males from Talybont (0.496), and was chosen in order for the mean of the zygotes to be about the same as that observed among the autumn eggs. It was assumed that in the 2B males the B chromosomes paired in 67% of the cells, the value reported by HEWITT(1973a), and that in the 2B females they paired in 25% of the cells, again the value reported by HEWITT(1976), and that the paired B chromosomes segregated regularly. As may be seen from the last comparison in Table 4, the frequencies of the various karyotypes among the zygotes calculated in this way were quite similar to those found among the autumn eggs and thus, it is no longer necessary to assume that strong selection against the OB embryos operated between the laying and the collection of the autumn eggs. Moreover, when these zygotes were used as a basis for determining the relative survival of the various karyotypes from zygotes to the adults, the B chromosome again appeared to reduce the survival of all the individuals carrying it (the last row in Table 4). Thus, it would appear that the evidence that in this population the B chromosome is “heterotic,” is quite weak, and that it may actually be “parasitic.” The above analysis and discussion indi- cates that the possibility cannot be ruled out that a B chromosome possessing an accumulation mechanism might be “heterotic,” especially if the accumulation is small and/or the frequency of individuals with two or more B chromosomes is fairly high. So far, however, all B chromosomes with an accumulation mechanism for which data on fitness are available appear to be of the “parasitic” type. Are the “parasitic” B chromosomes adaptive? The finding that a B chromosome with an accumulation mechanism reduced the fitness of all the individuals carrying it led KIMURAand KYANO(1961), as quoted earlier, and NUR (1969) to conclude that it was maintained because of its accumulation mechanism. Several authors, however, criticized this conclusion (see JONES1975). For example, WHITE(1973) questioned whether B chro- “PARASITIC” B CHROMOSOME 51 1 mosomes could have persisted “through all the vicissitudes of inter-deme selection unless they possessed some adaptive significance” (p. 327) and appeared unwilling to accept the possibility “that the genotype of the species is unable to evolve in a way that would get rid of the ‘parasitic’ supernumeraries, or render them innocuous” (p. 315). Similarly, REESand AYONOADU(1973) pointed to the known effect of some B chromosomes on the amount of recombination of the regular chromosomes . . . and, following DARLINGTON( 1956), suggested that the B chromosome might be maintained through their advantage to the population as a whole, even though they might be harmful to the individuals carrying them. REES and AYONOADU,however, did not present any evidence indicating either that the effect of the B chromosomes on recombination was indeed adaptive, or that it would have been sufficient to counteract a strong selection against them. Thus, at present, it would appear that the question of the possible adaptive role of “parasitic” B chromosomes could not be resolved without a better under- standing of the relative strengths of intra- and inter-deme selection. However, since WHITE’S(1973) and REES and ATONOADU’Sideas about the adaptive role of B chromosomes should apply to all B chromosomes, even those lacking an accumulation mechanism, the most convincing evidence for the role of inter- deme selection in the maintenance of “parasitic” B chromosomes would be the discovery of a “parasitic” B chromosome that lacks an accumulation mechanism. I would like to thank DR. CONRADA. ISTOCKfor commenting on the manuscript.

NOTE ADDED IN PROOF A new sample of 128 males was collected between August 25 and September 19, 1977, from the same area as the 1974 sample discussed earlier. Eight of the males possessed one metacentric B Chromosome, two possessed acrocentric B chromosomes, and one possessed two metacentric B chromosomes. The B chromosomes of the latter differed from the metacentric B chromosomes of the other males in being isopycnotic during prophase I of meiosis (rather than heteropycnotic) and thus may represent a new type of B chromosome. The frequency of males with metacentric B chromosomes (7%, including the male with two metacentric B chromosomes) in the 1977 sample is significantly lower than in the 1974 sample of males (15.2%) (xz= 4.476, df = 1, p < 0.05). Thus, the fiequency of this B chromosome is apparently not as constant from year to year a? ha4 been assumd earlier. The resuits of the new sample, hcwever, provide further sup- port for the main conclusions reached earlier, namely that the frequency of the metacentric B chromoscme does not increase from generation to generation in spit? of its high transmission by the females and that its failure to increase is due, at least in part, to selection against 1B individuals.

LITERATURE CITED

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