Heredity72 (1994) 155—1 62 Received 14 June 1.993 Genetical Society of Great Britain

Reproductive isolation between two species of flour beetles, Tribolium castaneum and T. freemani: variation within and among geographical populations of T castaneum

MICHAEL J. WADE* & NORMAN A. JOHNSON Department of Ecology and , 1101 E. 57th Street, , Chicago, IL 60637,USA

Tribolium casraneum and T freernani produce sterile hybrid progeny in reciprocal crosses. The reciprocal crosses differ significantly in the mean numbers of progeny, progeny sex ratios, hybrid male body size and male antennal and leg morphologies. These results suggest an effect of either the X chromosome or the cytoplasm on characteristics of F1 hybrids. In contrast, large X chromosome effects on morphological traits are not usually oberved in interspecific crosses among drosophilid flies. We also report large, significant differences in progeny numbers, body mass and degree of female bias in sex ratio between different geographic strains of T castaneum when mated in reciprocal crosses with T freemani. Sex ratio bias also varies significantly among matings within geographic strains of T castaneum. When T castaneum males are mated with T freemani females, but not in the reciprocal cross, the F1 sex ratio is female biased, uncorrelated with family size and ranges from 57.14 per cent to 72.23 per cent female, depending on the geographic strain of the T castaneum male.

Keywords: hybridinviability, genetic variation,morphology, reproductive isolation, , Tribolium

sex that is most adversely affected. This rule applies to Introduction taxa in which males are the heterogametic sex (includ- Dobzhansky(1937),Mayr(1963) and others (Coyne et ing beetles) as well as in cases wherein females are the al., 1988; Coyne, 1992b; Wu & Davis, 1993) have heterogametic sex. Recently a number of authors ('Mi, argued that the evolution of genetic reproductive isola- 1992;Wu & Davis, 1993; Orr, 1993; Partridge, 1993) tion is critical to the formation of new species. One have come to the conclusion that Haldane's rule is type of reproductive isolation is hybrid inviability. probably a composite phenomenon with different Although there is a burgeoning volume of information explanations applicable to different aspects (sterility vs. about the patterns and the genetics of hybrid inviability inviability, heterogametic males vs. heterogametic in drosophilid flies (Crow, 1942; Watanabe, 1979; females) of the rule. Although no complete inviability Bock, 1984; Coyne & Orr, 1989a,b; Hutter et al., of either sex has been observed in the hybrids of 1990; Sawamura et al., 1993a,b); Orr, 1993; Wu & crosses between T castaneum and T freemani Davis, 1993), little is known about the genetic basis of (Brownlee & Sokoloff, 1988), it is not known whether hybrid inviability in other taxa, such as the notably there are more subtle differences in the viabilities of the speciose Coleoptera. two sexes in the hybrids. Coyne & Orr(1989a) have directed attention to two The second empirical generalization is the large empirical generalizations about the genetics of post- effect of the X chromosome. In most cases of post- mating reproductive isolation. The first of these is mating reproductive isolation (most strongly supported Haldane's rule (Haldane, 1922) which states that in for cases of hybrid sterility), the X chromosome has a crosses between different animal races or species, if disproportionate effect. This rule does not seem to there is sterility or inviability in one but not the other apply to traits involved in premating (behavioural) sex of the F1 hybrids, it is most often the heterogametic reproductive isolation (Coyne, 1989, 1992a, 1993) or to morphological traits (Coyne, 1985; Charlesworth et *Correspondence al., 1987; Coyne & Orr, 1989a; Spicer, 1991; but see 156 M. J.WADE & N. A. JOHNSON

Templeton, 1977). These generalizations, however, product of the forces of and/or have been based almost exclusively on data from random genetic drift (Dobzhansky, 1937; Mayr, 1963; Drosophila genetic studies. We can infer whether a Carson, 1975; Templeton, 1980; Nei et a!., 1983; large X effect exists from a comparison of the male Charlesworth et at., 1987). In highly subdivided popu- morphologies of the F1 from reciprocal crosses (which lations, local adaptations that might affect the degree of differ only in the source of the X chromosome, Y reproductive isolation with another species will spread chromosome and cytoplasm). slowly through the species by migration if globally Tribolium castaneum is a human commensal adaptive but the extent of their spread could be quite species of flour beetle with a worldwide distribution restricted if only locally adaptive. In addition, if found in a variety of stored products (Sokoloff, 1974). random genetic drift were responsible for the repro- I freemani is two to three times the mass of I ductive isolation between species (see Nei et at., 1983), castaneum but otherwise morphologically and karyo- we would also expect considerable heterogeneity typically similar (eight pairs of autosomes, females X)(, among populations from limited migration within males XY) (Hinton, 1948; Sokoloff, 1974; Nakakita et species early in the speciation process. Thus, in the a!., 1981; Brownlee & Sokoloff, 1988). Certain geo- early stages of speciation there might be considerable graphic and laboratory genetic strains of I castaneum intraspecific genetic variation for pre- and postmating produce large numbers of F1 progeny in reciprocal reproductive isolation, not only between reciprocal hybridizations with T freemani. In many cases, the pro- crosses among species but also, or even especially, ductivity of F1 crosses is comparable to that found in among populations within a species. intraspecific crosses (see below). Nearly all the hybrids In this study of the species pair, T castaneum and T of both sexes are sterile or, at best, a few (less than 1.33 freemani, we report finding such intraspecific variation per cent) are very weakly fertile (producing fewer than in hybrid progeny numbers, the degree of female sex five progeny) (Brownlee & Sokoloff, 1988; Wade et al., ratio bias and male body size at three levels: (1) varia- 1994; see below). tion between reciprocal crosses, (2) variation among These two species do not exhibit any detectable different geographic and laboratory strains of I premating reproductive isolation (Wade et at., 1993), castaneum, and (3) variation among individuals or consistent with the indiscriminate nature of mating families within strains. within Tribolium (Park, 1933; Sokoloff, 1974). Some kind of postcopulatory but prezygotic reproductive Materialsand methods isolation exists between these two species (Wade et al., 1994): when females of either species mate with both Virginmale and female I freemani were mated conspecific and heterospecific males, almost all off- individually to virgins from one of several strains of T spring are sired by the conspecific male (i.e. few castaneum. The following five strains of I castaneum hybrids are produced) (Wade et at., 1994). were used: (A) c-SM: an outbred mixture of the four Here, we report variation in the numbers of F1 classic 'Park' strains (Park et a!., 1964; Wade, 1977), progeny, progeny sex ratio and F1 body size within and (B) c-pI: a strain segregating the pearl eye mutation between geographical strains of T castaneum when obtained from Dr R. Beeman at the USDA Grain reciprocally hybridized with I freemani. The empirical Research Marketing Laboratory at Lawrence, Kansas, literature on the genetic basis of reproductive isolation (C) c-Yugo: a wild strain collected in 1979 in the former often ignores variation in the degree of reproductive Yugoslavia, now Croatia, (D) c-BS: a wild strain col- isolation between interbreeding species that may exist lected in 1987 in Naperville, Illinois, and (E) c-Inf: a within or among populations of either species. Conse- wild strain collected in 1979 in Infantes, Spain. The quently, there are few studies of intraspecific variation c-SM strain is an outbred laboratory strain and the last in features of interspecific hybridization. Those that do three wild strains were derived from collections con- exist generally focus on variation in major genes, such sisting of at least 50 adults each. The T freetnani strain as hmr (hybrid male rescue) which rescues inviable F1 used was the standard stock, collected in 1978 (Naka- male progeny in crosses between Drosophila melano- kita et a!., 1981) from a shipment of grain to Japan gaster and D. simulans (see Watanabe, 1979; Hutter et from Chile. at., 1990; Sawamura et at., 1 993a,b). In contrast, most For each of the above strains, except c-BS and c-mi, scenarios for the evolution of genetic reproductive iso- 25—28 replicates of reciprocal crosses were established lation between species implicitly assume heterogeneity in an 8 dram vial containing 8 g of standard medium within populations. They postulate the adventitious (95 per cent sifted stoneground flour, 5 per cent yeast; fixation within each of two species of genes affecting see Wade, 1977). For c-BS, 15—17 reciprocal crosses reproductive isolation occurring as an incidental by- were set up but for c-Inf only nine matings of males to VARIATION IN REPRODUCTIVE ISOLATION 157

T freemani females were set up. All pairs were main- The reciprocal cross, F X c-SM, experienced more tained in a darkened incubator at 29°C with 70 per failed matings (9 of 25) and nearly 60 per cent lower cent relative humidity for 55 days. We note that as each productivity per cross (Table 1). The progeny sex ratio, family was raised in just one vial, the among-family 52.02 per cent females, was not significantly different effects described in Results below may occur from vial from unity (G=0.607, 1 d.f.) although it did differ effects rather than from genetic differences. (In related from that of the reciprocal cross (x2= 11.029,1 d.f., work, repeated observations on single females of both P<0.001). The FXc-SM families were not signifi- species indicate significant family as opposed to vial cantly heterogeneous in sex ratio (G-heterogen- effects (R. Jones and M. J. Wade, unpublished data).) eity=21.003, 16 d.f.). Only 6 male and 6 females of After 55 days, all cultures were sifted and the off- 371 total (3.2 per cent) exhibited the friability of spring were censused, separated by sex and weighed to antennal segments commonly observed in males of the the nearest 0.01 mg on a Mettler AE163 balance. At reciprocal cross (see above). this time, morphological abnormalities of the antennae Of 300 c-SM x F female hybrids tested, only 3 and legs of individual beetles were also noted. Statisti- females were weakly fertile, each producing fewer than cal tests were performed using the STATISTICA STATSOFT 3 larvae in backcrosses to either T castaneum or T computer software package. freemani males. None of the 120 reciprocal hybrid We will discuss the results of the interspecific crosses females tested was fertile. Overall, 0.71 per cent (3 of first by geographic strain of T castaneum and then 420) of all hybrid females were weakly fertile. review patterns common to all geographic strains. The Hybrid males of the cross F male X c-SM female had two laboratory strains, the outbred c-SM and the a mean weight of 2.944 mg with a standard error of mutant stock, c-pl, will be discussed first, followed by 0.03832 mg (N= 148). This was much closer to the data for the three wild stocks, c-Yugo (Zagreb), c-BS mean of the weights of pure species c-SM (mean weight (Naperville, IL) and c-Inf(Tnfantes, Spain). of c-SM males 2.370 mg, standard error= 0.04532 mg, N= 23; mean of weight of c-SM females 2.493 mg, Results standard error 0.03392 mg, N= 24) than it was to the mean of pure species T freemani (male mean =6.013 Wefirst describe the major features of productivity and mg, standard error=0.1181 mg, N=22; female sex ratio of the reciprocal crosses of each strain of T mean=6.106 mg, standard error=0.1645 mg, castaneum when hybridized to T freemani. N= 16). There was significant among-family variation for male weight in this cross (ANOVA, 14 among d.f., 133 error d.f., F= 16.387, P° 0.000 1). Hybrid females c-SMhybridizations with T. freemani of this cross were heavier, averaging 3.707 mg with a Crossesbetween L castaneum males and T freemani standard error of 0.04208 mg (N 173). There was females (c-SM >< F) are less likely to fail to produce also significant heterogeneity among families for hybrid progeny (3 of 25 vs. 9 of 25 in reciprocal matings, female weight (ANOVA, 14 among d.f., 158 error =3.95,0.025 < P< 0.050) and produce significantly F=6.771, P°(<0.0001). The male and female mean more progeny than the reciprocal cross, FX c-SM weights across families were highly correlated (Spear- (Table 1). The sex ratio of the c-SM X F progenies man's rank order correlation, r =0.878,P< 0.0001). (62.75 per cent female) differed significantly from unity In the reciprocal cross (c-SM male X F female), the (G-test, 1 d.f., G=42.547, P<0.001). The progenies hybrid females weighed on average 4.412 mg with a of different families also varied significantly in sex standard error of 0.03246 mg (N= 182). There was ratio from one another (G-heterogeneity = significant heterogeneity among families for female 51.60,21 d.f., P<0.001), ranging from 87 per weight in this cross (ANOVA, 18 among d.f., 163 error cent females (33 females of 38) to 36 per cent females d.f., F= 2.35 1, P= 0.0250). From this cross, there were (9 females of 25). We observed no correlation between too few males (19) which survived at the time of family size at day 55 and sex ratio (r= —0.043,17 d.f.). weighing to warrant inclusion in these data. We noted that many male, but not female, hybrids in this cross were missing several of the terminal antennal hybridizations with 1. freemani segments or the entire antennae, which appeared to be c-p/ broken off. Complete data were gathered from four Crossesbetween c-pl males and Tfreemanifemales (c- families in which 15 of 39 males or 38 per cent p1 x F) fail to produce progeny more often than the exhibited this condition but none of 86 females did so. reciprocal cross, FX c-pl (4 of 25 vs. 0 of 25 matings, (See c-BS hybridization with T freemani for a complete chi-square 4.34, 0.025

Table 1 Mean and standard error of the numbers of male and female progeny produced in crosses between T eastaneum and Tfreemani

C-male x F-female F-male XC-female

Strain of Male Female Male Female T castaneum Mean (S.E.) Mean (S.F.) Mean (SE.) Mean(SE.)

c-SM 9.6 (1.48) 16.2 (2.15) 7.1 (1.66) 7.7 (1.69) Total 25.9(3.24) 14.8(3.18) c-pl 7.9 (1.52) 20.6 (1.40) 37.7 (3.31) 42.7 (4.12) Total 28.5 (3.38) 80.4 (6.89) c-Yugo 13.0(1.19) 17.3 (1.39) 28.8 (3.90) 30.7 (3.97) Total 30.3 (2.06) 66.6 (6.89) c-BS 7.2(1.49) 10.7 (2.31) 14.9 (3.23) 14.6 (3.42) Total 17.9 (3.47) 29.5 (6.53) c-Inf 8.0 (1.86) 19.0 (3.30) — Total 27.0 (4.04) —

c-pl x F progenies averaged 72.23 per cent female, the d.f., 316 error d.f., F= 14.974, P40.0001). Hybrid largest average difference of any strain from a sex ratio females resulting from this cross weighed on average of unity (G-test, 1 d.f., G= 145.993, P40.001). These 3.857 mg with a standard error of 0.02084 mg progenies were also extremely heterogeneous in sex (N= 280). The weights of these females also were sig- ratio (G-heterogeneity 93.45, 20 d.f., P40.001), nificantly heterogeneous among families (ANOVA, 4 ranging from 97 per cent females (32 females of 33) to among d.f., 275 error d.f., F== 11.632, P40.001). 47 per cent females (25 females of 47). There was no There were not enough families, for which we obtained significant correlation between family size at day 55 data on both male and female weights, to test for a and sex ratio (r= + 0.275, 19 d.f.). Lastly, we noted correlation. No measurements were made on the that many of the male hybrids in this cross, but never offspring from the reciprocal cross. females, had several of the distal antennal segments and the tarsi broken off. c- i The reciprocal cross, Fx c-pl, experienced no failed Yugo hybridizations with T. freeman matings at all (0 of 25) and nearly a threefold higher Crossesbetween T castaneum males from Yugoslavia productivity per cross (Table 1). The average progeny and Tfreemanifemales (c-Yugo >

F x c-Yugo families were also significantly heterogene- Of the 200 female hybrids tested for fertility by ous in sex ratio (G-heterogeneity=42.98, 25 d.f.), backcrossing to the parental species, only 1 Fx c-BS ranging from 66.3 per cent females (69 of 104) to 31.0 female produced a single offspring when crossed to a T per cent females (13 of 42). castaneum c-SM male. None of the 770 mass mated The 150 c-YugoXF hybrid females were tested by hybrids produced progeny, suggesting that the males backcrossing to males of the parental species. In two are also sterile. cases, hybrid females backcrossed to T castaneum FX c-BS hybrid females had a mean weight of 3.452 males were weakly fertile, each producing fewer than mg with a standard error of 0.02173 mg (N=239). five larvae. In backscrosses of 150 Fx c-Yugo female There was significant variation in weight among ten hybrids, two females were weakly fertile when mated randomly selected families (ANOVA, 9 among d.f., with T freemani males. 229 error d.f., F=7.521, P40.0001). Hybrid males of The F Xc-Yugohybrid males had a mean weight of this same cross weighed on average 2.608 mg with a 2.996 mg with a standard error of 0.03772 mg standard error of 0.01908 mg (N= 249) but no signifi- (N= 152) whereas their sisters had a mean weight of cant variation among eight different families was found 3.844 mg with a standard error of 0.03143 mg (ANOVA, 7 among d.f., 241 error d.f., F= 1.352, (N= 296). In the reciprocal cross (c-Yugo XF),hybrid P>0.05). males had a mean weight of 4.7452 mg and a standard In the reciprocal cross (c-BS >

Table 2 Sex ratio of F! hybrids between different strains of Table 3 Summary of weights of hybrid males and females T castaneurn and T freernani 7: castaneurn strain C-male x F-female F-male x C-female Strain of Female (N) Female (N) c-SM c-pl c-Yugo c-BS Tcastaneurn (%) (%) Fx ct c-SM 62.75 (647)1: 52.02(371) Males 2.9441: 2.900 2.996 2.608 c-pl 72.23 (713)1 53.16 (2009)1: 0.03832 0.01783 0.03772 0.01908 c-Yugo 57.14 (847)1 51.59(1605)1: (148) (322) (152) (249) c-BS 59.85 (269)1: 49.50 (501) Females 3.707 3.852 3.844 3.452 c-Inf — 70.40 (243)1: 0.04208 0.02084 0.03143 0.02173 1: Indicates significant female bias and significant (173) (280) (296) (239) heterogeneity in sex ratio among families. cxF 1: Indicates significant heterogeneity among families in sex Males — — 4.742 4.33 1 ratio. 0.04975 0.06777 (143) (105) Females 4.412 — 3.876 3.850 0.03246 0.02225 0.03352 In the reciprocal cross to T freemani males, only the (182) (316) (101) laboratory mutant strain, c-pl, exhibited a small but F-c males — — 1.746 1.723 significant female bias in hybrid sex ratio. It was not as 0.705 — 0.032 0.398 female-biased as any of the reciprocal crosses to T F-c females freemani females. The sex ratios were not heterogene- tFX c represents the cross of a T freernaniinale to a T ous among strains, however (G-heterogeneity =2.440, castaneurn female; c XFis the reciprocal cross. 3 d.f., not significant). Both the c-Yugo and c-pl strains 1:Upper number is the mean weight in mg, middle number is exhibited very significant among-family variation in sex the S.E.M. The number tested is in parentheses. ratio. However, in both strains, the most heavily §F-c males and F-c females represents the difference in the female-biased families exhibited sex ratios below the weights in hybrid males and females, respectively, from the average sex ratio observed in most of the reciprocal cross in which T- freernani is the female and the cross in crosses. which T castaneurn is the female. The relative size of male and female F1 hybrids dif- fered between the reciprocal crosses (summarized in Table 3). Hybrid males were generally 15 per cent larger than hybrid females when T freernani was the Discussion mother but males were 25 per cent smaller than females when T castaneum was the mother. Hybrid Ourobservations are consistent with the inviability male weight in the former cross was 1.65 times as great aspect of Haldane's rule in that when one sex in the F1 as in the latter. Females of both pure species weigh up is rarer it is the males (which are heterogametic in to 10 per cent more than males as is typical of species Tribolium). For each of the five strains of T castaneum in the genus (see above; Sokoloff, 1974). The sex dif- males mated to T freemani females and in two of four ference in weight in the hybrids is greater than that reciprocal crosses, we observed significant heterogen- observed within the parent species and, in one of the eity among families in progeny sex ratios. In some interspecific crosses, the sexual dimorphism in size is cases, the variation ranged from a complete absence of reversed. males to equality of sex ratio. However, no correlation In three strains, c-SM, c-pl and c-BS, we noted the within any geographical or laboratory strain was seen integrity of antennal segments in males and females of between family size at day 55 and the degree of sex both reciprocal crosses. When crossed to T freemani ratio bias in favour of females. A negative correlation females, male but not female hybrids exhibited fragile between family size and degree of female bias might be or friable antennal segments. In both c-SM and c-BS, expected if male mortality were responsible for the the incidence is approximately 30—40 per cent of all rarity of males. With the number of crosses observed, hybrid males. This condition was not observed to any we should have been able to detect a correlation of significant degree in either sex in the reciprocal crosses 0.45 at the 0.05 level; the observed correlations were of females of these strains to T freemani males. all smaller (see above). Of course, mortality observa- VARIATION IN REPRODUCTIVE ISOLATION 161

tions comparing the numbers of adults at day 55 to the evidence that there is variation for hybrid inviability. numbers of eggs laid or larvae hatched would be more Hybrid inviability as a form of postmating reproductive conclusive. (Preliminary observations on work in pro- isolation is generally thought not to be adaptive in itself gress suggest that the rarity of males is due to but the by-product of different adaptive or neutral mortality.) changes evolving within each of two nascent species We have observed that morphological abnormalities (Dobzhansky, 1937; Mayr, 1963; Charlesworth et at., (for instance, broken distal antennal segments and 1987; Coyne, 1992b; Wu & Davis, 1993). Coyne deformities of the distal leg segments) occur more often (1974), however, has argued that hybrid inviability in hybrids of the heterogametic sex (male) than in the itself may be an adaptive trait if the hybrids are sterile homogametic sex. Thus we have observed a morpho- and there is parental care. He argues that, if the hybrids logical equivalent of Haldane's rule. are sterile (either physiologically or behaviourally), it Data from Drosophila hybridizations also strongly would be costlier to provide parental care than to have suggest that hybrid male sterility evolves much faster them be inviable. We believe that competition occur- than hybrid female sterility or hybrid inviability (Coyne ring within families may substitute for the requirement & Orr, 1989b; Wu, 1992; Wu & Davis, 1993; Johnson of parental care in Coyne's argument. That is, if & Wu, 1993). Our data are consistent with a faster rate individuals produce both sterile hybrids and pure- of evolution to hybrid sterility compared with that to species progeny and the different types of progeny hybrid inviability as most hybrids of both sexes are compete for resources, selection between families sterile and none approaches normal fertility. In would favour the (early) inviability of hybrids. contrast, the hybrids from most of the crosses appear These conditions may be met in natural populations to have intermediate to normal viability. of Tribolium. First, almost all of the hybrids between T The large difference in weights between reciprocal freemani and T castaneum are completely sterile F1 males suggests that the X chromosome, the Y (Brownlee & Sokoloff, 1988; Wade et al., 1994; this chromosome and/or the cytoplasm have a dispropor- study). Secondly, there is evidence that the densities of tionate effect on this morphological trait. Because the Tribolium populations may be regulated at a small- reciprocal F1 females, who also have different sources spatial scale giving rise to within-family competition of their cytoplasm, do not demonstrate as large a dif- (Park et al., 1964; Sokoloff, 1974; McCauley & Wade, ference in weights, it is more probable that the X or the 1980). Wade & Beeman (1994) have suggested, on Y chromosomes rather than the cytoplasm is respon- theoretical grounds, that the spread of the Medea sible for the majority of the weight difference. Further- selfish element present in Tribolium, is facilitated by more, the effect reverses the sexual dimorphism in within-family competition. Likewise, we suggest that body size, typical for both species, wherein males are the within-family competition may have facilitated the smaller than females. We cannot distinguish between evolution of hybrid inviability. Here we have shown an X effect and a Y effect but as the Y chromosome is that significant variation exists among populations of T generally genetically depauperate, it is more likely that castaneum as well as among families within populations it is the X chromosome that is responsible for these for this potentially adaptive trait. differences. Moreover, it is the larger of the F1 male hybrids (c male x F female) that receives its Y chromo- Acknowledgements some from T castaneum, the smaller of the two parental species; thus the size differences are probably Wethank Nancy Chang, Michael McNaughton and not due to a Y effect. Ora Lucas for technical assistance. We thank Jerry The deformities of the distal segments of the male Coyne, John Kelly, Mohamed Noor, Mike Palopoli and antennae and legs observed in this same cross but not Chung-I Wu for discussion of and/or comments on the in the reciprocal follow the same pattern as male manuscript. This work has been supported by a National Institutes of Health grant (GM 22523) to weight. These reciprocal (apparent large X) effects for morphology contrast with the results from Drosophila M.J.W. hybridizations. We suggest that the difference may be due to different genetic architectures being responsible References for the traits observed here, body size and antennal BOCK,I. R. 1984. Interspecific hybridizations in the genus integrity, compared with the traits studied in Droso- Drosophila. Evol. Biol., 18, 41—70. phila hybridizations (e.g. genital arch shape (Coyne, BROWNLEE, A. ANDSOKOLOFF,A.1988.Transmission of Tn- 1985) and colour band size (Spicer, 1991)). bolium castaneum (Herbst) mutants to T castaneum-T The large differences in the productivities among freemani Hinton hybrids (Coleoptera: Tenehrionidae). J. and within strains in the interspecific crosses is Stored Prod Res., 24, 145—150. 162 M. J. WADE & N. A. JOHNSON

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