Industrial and non-industrial melanism in the peppered moth, Bistort betularia (L.)

Abstract. 1. Estimates of the relative crypsis of the melanic and typical morphs of B.betularia have been made at fifty-two sites in southern England and south Wales and these estimates were compared with melanic frequencies in samples from these sites.
2. The decrease from east to west in the frequency of the melanic form, carbonaria , and increase in the frequency of a second melanic form, insularia , across the southern part of England and Wales is largely independent of changes in smoke concentration, sulphur dioxide concentration or relative crypsis.
3. When all the available information on the distribution of the melanics of B.betularia is considered, sulphur dioxide concentration is correlated with the geographic variation in carbonaria frequency. This, together with evidence of the close relationship between carbonaria crypsis and sulphur dioxide levels, indicates that outside southern England and south Wales, either selective predation, or some direct selective effect of pollutants, is of major importance in determining the variation in carbonaria frequency.
4. Some features of the pattern of the spread of carbonaria in England and Wales during the last century give reasons for expecting non-industrial selective factors to be of greater importance in determining the distribution of melanics of B.betularia in southern England and south Wales than in the rest of Britain.     

Fitness of insularia morphs of the peppered moth Biston betularia

In recent years the industrial melanic carbonaria morph in the moth Biston betularia (L.) has decreased rapidly in frequency in Britain as air pollution has decreased. The intermediate melanic insularia has shown a variable response. We have estimated the fitness of insularia, compared with the other two morphs, for several data sets. As a rule its fitness lies between that of carbonaria and typical, but nearer to typical and sometimes very close to it. The intermediate position is expected if fitness relates directly to phenotype. The results suggest that insularia may continue polymorphic while carbonaria is likely to disappear. The past high frequency of insularia in South Wales may have been due to an initial increase in insularia frequency before carbonaria reached the region. Differences in dynamics of frequency change in insularia and carbonaria are evidence against induction, which has sometimes been invoked to explain the spread of melanism in this species.  © 2004 The Linnean Society of London, Biological Journal of the Linnean Society , 2004, 82 , 359–366.     

Peppered moth revisited: analysis of recent decreases in melanic frequency and predictions for the future

In a recent review article by Mani published in 1990, it was shown that the changes in the frequency of the morph carbonaria can be explained through a migration-selection model and the model is capable of reproducing the decrease in the carbonaria frequency since the enactment of Clean Air Acts. In this paper new data both for carbonaria and for insularia up to 1990 for West Kirby in NW England and for Cambridge are presented. In the earlier review, the data for West Kirby extended to 1987 and for Cambridge to 1985. Data not previously published for the carbonaria and the insularia frequencies at sites in Northwood in Middlesex, Egham in Surrey and Ringwood in Hampshire are also presented. A migration-selection model was used to predict the change in the frequencies of the melanic morphs up to the year 2010. All the data are compared with the predictions of the model. It is encouraging that the data follow the theoretical predictions.     

A decline of melanism in the peppered moth Biston betularia in The Netherlands

Melanic forms of the peppered moth Biston betularia were well established in The Netherlands by the end of the 19th century, indeed the first records of the black carbonaria form in 1867 are only about 20 years later than in England. Analysis of extensive sampling data collected by B. J. Lempke for a period of several years beginning in 1969 shows that carbonaria was at a frequency of about 60 to 70% in most of the country where epiphyte communities on trees were reduced due to the effects of air pollution. The pale typica and the three intermediate insularia forms were each at similar, low frequencies. Only in the extreme north and south-east of The Netherlands where epiphyte floras were richer was carbonaria at a lower frequency of less than 40%. Samples collected from seven localities in 1988 show that carbonaria has dramatically declined to a frequency of less than 10%. In contrast to England, the fully black form is being replaced not only by typica but also by the darkest of the insularia phenotypes. The decline in melanism coincides with a period of decreasing levels of sulphur dioxide and of increasing species diversity of lichens on trees.     

The postfire discomycete Geopyxis carbonaria (Ascomycota) is a biotrophic root associate with Norway spruce (Picea abies) in nature

The hypothesis that the postfire discomycete Geopyxis carbonaria (Ascomycota, Pezizales, Pyronemataceae) has a biotrophic association with roots of Norway spruce ( Picea abies ) in nature was tested by isolation of fungal strains from fresh, brown, smooth mycorrhiza-like root tips of Norway spruce collected from below the depth of detrimental heat penetration in a postfire site. The morphology of seven culture isolates originating from the smooth mycorrhiza-like root tips of two different spruce trees was congruent with the morphology of axenic culture isolates obtained from ascospores of G. carbonaria . DNA sequences of the nuclear ribosomal internal transcribed spacers ITS1 and ITS2 from these root-derived cultures and the ascosporic G. carbonaria culture isolates were found to be identical, further supporting the conclusion that the isolates were conspecific. The extensive ascocarp and ascospore formation of G. carbonaria which succeeds a forest fire may be explained in terms of a fungal escape from a moribund tree associate. Possible ecological adaptations of G. carbonaria to the pre- and postfire community are discussed.     

Antibiosis and tolerance to five species of spittlebug (Homoptera: Cercopidae) in Brachiaria spp.: implications for breeding for resistance

Several genera and species of spittlebugs (Homoptera: Cercopidae) are economic pests of Brachiaria spp. grasses in tropical America. To support current breeding programs aimed at obtaining multiple spittlebug resistance, we undertook a series of studies on antibiosis and tolerance as possible mechanisms of resistance to five major spittlebug species affecting Brachiaria spp. in Colombia: Aeneolamia varia (F.), Aeneolamia reducta (Lallemand), Zulia carbonaria (Lallemand), Zulia pubescens (F.), and Mahanarva trifissa (Jacobi). Four host genotypes, well known for their reaction to A. varia attack, were used to compare their resistance to other spittlebug species: CIAT 0654 and CIAT 0606 (susceptible) and CIAT 6294 and CIAT 36062 (resistant). CIAT 0654 and CIAT 36062 were used in antibiosis studies. Tolerance studies were conducted with CIAT 0654, CIAT 6294, and CIAT 36062. Sixty-five hybrid-derived clones were used to identify levels of multiple resistance to three spittlebug species. The levels of antibiosis resistance in CIAT 36062 clearly differed by spittlebug species and were classified as follows: very high for M. trifissa, high for A. varia and A. reducta, moderate for Z. pubescens, and absent for Z. carbonaria. Our results suggest the presence of true tolerance to Z. carbonaria in CIAT 6294 and CIAT 36062, true tolerance to Z. pubescens in CIAT 6294 and a combination of tolerance and antibiosis as mechanisms of resistance to Z. pubescens in CIAT 36062. Of the 65 hybrid clones tested with A. varia, A. reducta, and Z. carbonaria, 15 combined resistance to two species and three showed antibiosis resistance to all three spittlebug species.     

Nest architecture and genetic differentiation in a species complex of Australian stingless bees

We investigated the taxonomic significance of nest shape and its putative role in speciation in Trigona (Heterotrigona) carbonaria and T. (H.) hockingsi, two sibling species of stingless bee species from eastern Australia. These species are primarily distinguished by their nest architecture, as in all other respects they are nearly identical. We genotyped 130 colonies from six locations in Queensland at 13 microsatellite loci together with 106 additional colonies from six other Indo-Pacific Trigona species. Whether they were present in allopatry or in sympatry, colonies that displayed the T. carbonaria or the T. hockingsi nest architecture could be unambiguously differentiated at the genetic level. However, T. hockingsi colonies were classifiable into two highly differentiated paraphyletic and geographically separate populations, one in northern and one in southern Queensland. These two populations probably belong to two distinct species, T. hockingsi and T. davenporti nov. sp. Our results suggest that nest architecture characters are relevant but not sufficient criteria to identify species in this group. Consequently, modifications of nest architecture are probably not of prime importance in the speciation process of Australian stingless bees, although nest architecture differences probably result from relatively simple mechanisms. The rare interspecific hybrid colonies detected did not display a nest with an intermediate form between T. hockingsi and T. carbonaria.     

The rise and fall of the Carbonaria form of the peppered moth

The evidence for change in frequency of the melanic carbonaria morph in the peppered moth Biston betularia (L.) (Lepidoptera: Geometridae) in England and Wales is reviewed. At mid-20th century a steep cline of melanic phenotype frequency running from the north of Wales to the southern coast of England separated a region of 5% or less to west from 90% or more to northeast. By the 1980s the plateau of 90% frequency had contracted to northern England. The frequency has since continued to drop so that the maximum is now less than 50% and in most places below 10%. There have been similar declines in Europe and North America. Evidence from surveys and from two-point records shows the change to require 5% to 20% selection against the melanic. The melanic is more disadvantageous in regions where its frequency was initially high than in regions where it was low. Experiments to investigate predation by birds show a net advantage to carbonaria morphs in regions where typical frequencies were low at the time of the experiment, and a disadvantage where typical frequencies were high. This would be expected if environment and frequency were associated, and selective predation played a part in generating the association. The cryptic advantage of carbonaria was large in areas of heavy pollution where typical frequencies were 20% or less. The moth usually has a low density but is relatively highly mobile. The ability of present information to explain the patterns has been tested in simulations. They indicate a system under strong selection that has always been in a dynamic state without equilibria.     

Response of resistant and susceptible Brachiaria spp. genotypes to simultaneous infestation with multiple species of spittlebugs (Hemiptera: Cercopidae)

The response of one susceptible and three resistant Brachiaria spp. (Hemiptera: Cercopidae) genotypes to individual or combined attacks by nymphs of Aeneolamia varia (F.), Aeneolamia reducta (Lallemand), Zulia carbonaria (Lallemand), and Zulia pubescens (F.) was studied. We assessed the effect of infesting plants of the susceptible check BRX 44-02 and of the A. varia-resistant genotypes CIAT 6294 and CIAT 36062 with A. varia, Z. carbonaria, or Z. pubescens either alone or in two-species combinations. In a second trial, we studied the performance of BRX 44-02, CIAT 6294, and the multiple resistant clone SX01NO/0102 exposed to individual or combined attack by A. reducta and Z. carbonaria. In a third trial, we compared the response of BRX 44-02, CIAT 6294, and CIAT 36062 to individual A. varia, Z. carbonaria, or Z. pubescens attack as opposed to a combined three-species attack. Plant damage scores and percentage of nymphal survival were recorded in all three trials. Data on percentage of survival indicated that competition between and among spittlebug species occurs. However, we found no evidence of interaction between species competition and different levels of resistance to spittlebug. Rather, host genotype reactions conformed to previously known categories of resistance regardless of the presence of more than one spittlebug species. Resistance rather than competition seems to have been the overriding factor determining nymph survival and resistance expression (damage scores) in these experiments. Our results corroborate the need to develop brachiariagrass genotypes with multiple resistance to spittlebugs.     

Pre-adult viability differences of melanic Biston betularia (L.) (Lepidoptera)

Departures from mendelian ratios have been analysed in all published family data for the peppered moth, Biston betularia. No distinction was made between different insularia alleles; thus carbonaria is top dominant while insularia is dominant over typical. Viabilities of five of the six genotypes were calculated by a simple algebraic method; a maximum likelihood method gave viabilities for all six genotypes together with their standard errors. Carbonaria homozygotes (and possibly also insularia homozygotes) have enhanced viability while the viability of insularia/carbonaria heterozygotes is reduced. The significance of these findings is discussed in relation to phenotype frequencies found in the wild.     

Melanic frequencies in three species of moths in post industrial Britain

Changing patterns of morph frequency in three moth species ( Biston betularia , Odontoptera bidentata and Apocheima pilosaria ) have been investigated using data from the Rothamsted Insect Survey. All three exhibited industrial melanism during the period of high atmospheric pollution in Britain. Three historical and habitat types are compared, the old industrial north of England, rural Scotland, Wales and South-West England, and a southern English intermediate region of high human population density but generally low industrialization. Between 1974 and 1999 the carbonaria morph of B. betularia declined in frequency in the industrial region and is nearly absent from rural areas. It is the form which most closely tracks atmospheric change. It is shown that the insularia forms of B. betularia and the melanic morphs in the other two species have decreased in the industrial region, commencing later than carbonaria , but have maintained their presence and possibly reached equilibrium elsewhere. They may be non-industrial polymorphisms. B. betularia is rarer than the other species and all three species are at lower densities in industrial than in non-industrial regions. © 2002 The Linnean Society of London , Biological Journal of the Linnean Society , 2002, 75 , 475–482.     

Evolution in reverse: clean air and the peppered moth

Between 1848 and 1895 the melanic form f carbonaria of the peppered moth increased in Manchester from 0 to 98%. The reverse process is now occurring in one locality in Memyside where a year-to-year survey has shown that the pale form f. typica has increased from 6 to 30% between 1959 and 1984. Supporting information also comes from two studies in N Wales. The Clean Air Acts and the consequent decline in air pollution levels are probably responsible but there are many ill–understood problems, not the least being how, and the extent to which, non-visual selection operates.     

Evolutionary dynamics of declining melanism in the peppered moth in the Netherlands

Populations of Biston betularia in the region of The Netherlands around Leiden and Rotterdam were resampled. A comparison of three sets of data for 1969-1973, 1988 and 1999 enabled a further examination of declines in melanism. Unlike parallel changes for the black carbonaria form of this species in urban regions of Britain, those inThe Netherlands involve substantial changes in frequencies of at least two of the intermediate insularia morphs as well as an increase in the non-melanic typica morph. The darkest of the three insularia morphs has shown a transitory pulse of increased frequency in The Netherlands. The dynamics are discussed in relation to the history of air pollution and to straightforward predictions about selection.     

Sublethal effects of antibiosis resistance on the reproductive biology of two spittlebug (Hemiptera: Cercopidae) species affecting Brachiaria spp

Several greenhouse experiments were used to measure how high levels of antibiosis resistance to nymphs in two interspecific Brachiaria (brachiariagrass) hybrids affect life history parameters of the spittlebugs Aeneolamia varia (F.) and Zulia carbonaria (Lallemand), two of the most important spittlebug (Hemiptera: Cercopidae) species affecting Brachiaria production in Colombia. The A. varia-resistant hybrid CIAT 36062, the Z. carbonaria-resistant hybrid SX01NO/0102, and the susceptible accession CIAT 0654 were used to compare the effect of all possible combinations of food sources for nymphs and adults. Calculation of growth indexes showed a significant impact of antibiosis resistance on the biology of immature stages of both species. Median survival times of adults feeding on resistant genotypes did not differ from those recorded on the susceptible genotype, suggesting that factors responsible for high mortality of nymphs in the resistant hybrids did not affect adult survival. Rearing nymphs of A. varia on CIAT 36062 and of Z. carbonaria on SX01NO/0102 had deleterious sublethal effects on the reproductive biology of resulting adult females. It is concluded that high nymphal mortality and subsequent sublethal effects of nymphal antibiosis on adults should have a major impact on the demography of the two spittlebug species studied.     

Molecular characterization of hemoglobin alpha-D chains from Geochelone carbonaria and Geochelone denticulata land turtles

In order to help elucidate the evolution of alpha-globins, the complete cDNA and amino acid sequences of Geochelone carbonaria and Geochelone denticulata land turtles alpha-D chains have been described. In G. carbonaria, the cDNA is 539 bp with ATG start codon located at position 46, TGA stop codon at position 469 and AATAAA polyadenylation signal at position 520. In G. denticulata, the cDNA is 536 bp with ATG start codon located at position 46, TGA stop codon at position 469 and AATAAA polyadenylation signal at position 517. Both cDNAs codify 141 amino acid residues, differing from each other in only four amino acid residues. When comparing with human Hb alpha-chain, alterations in important regions can be noted: alpha110 Ala-Gly, alpha114 Pro-Gly, alpha117 Phe-Tyr and alpha122 His-Gln. There is a high homology between the amino acids of these turtles when compared with chicken alpha-D chains, progressively decreasing when compared with human, crocodile, snake, frog and fish alpha-chains. Phylogenetic analysis of alpha-D chains shows that those of turtles are closer to those of birds than to snakes and lizards.     

A new species of Syntretus Foerster (Hymenoptera: Braconidae: Euphorinae), a parasitoid of the stingless bee Trigona carbonaria Smith (Hymenoptera: Apidae: Meliponinae)

A new species of euphorine braconid, Syntretus trigonaphagus sp. nov., is described and illustrated. This is the first Australian Syntretus species to be described and the first record of braconids parasitising Trigona Jurine stingless bees. Egg-laying by adults and emergence of larvae from the host Trigona carbonaria is detailed. A 12S ribosomal rRNA gene fragment was sequenced to confirm the association of larvae and adults of the wasp parasitoid.     

Simple method for culture of peripheral blood lymphocytes of Testudinidae

We developed and optimized a simple, efficient and inexpensive method for in vitro culture of peripheral blood lymphocytes from the Brazilian tortoise Chelonoidis carbonaria (Testudinidae), testing various parameters, including culture medium, mitogen concentration, mitotic index, culture volume, incubation time, and mitotic arrest. Peripheral blood samples were obtained from the costal vein of four couples. The conditions that gave a good mitotic index were lymphocytes cultured at 37°C in minimum essential medium (7.5 mL), with phytohemagglutinin as a mitogen (0.375 mL), plus streptomycin/penicillin (0.1 mL), and an incubation period of 72 h. Mitotic arrest was induced by 2-h exposure to colchicine (0.1 mL), 70 h after establishing the culture. After mitotic arrest, the cells were hypotonized with 0.075 M KCl for 2 h and fixed with methanol/acetic acid (3:1). The non-banded mitotic chromosomes were visualized by Giemsa staining. The diploid chromosome number of C. carbonaria was found to be 52 in females and males, and sex chromosomes were not observed. We were able to culture peripheral blood lymphocytes of a Brazilian tortoise in vitro, for the preparation of mitotic chromosomes.     

A theoretical analysis of the morph frequency variation in the peppered moth over England and Wales

The migration-selection model for the spatial and temporal variation of morph frequencies of Biston betularia over England and Wales (Cook & Mani, 1980) has been extended to include effects due to non-visual selection. The parameters for non-visual selection were chosen from the recent determination by Mani (1980) and by Creed et al . (1980). The morph frequencies over England and Wales were obtained through computer simulation and the results were compared with data along the Manchester-Yorkshire, Central Wales-East Anglia and South Wales-London transects. Best fits to the data were obtained by using the non-visual selective values of Mani for carbonaria and modified values of Creed et al . for insularia . It is concluded that the observed polymorphism could be well explained through a balance of migration and visual and non-visual selections.     

Allelic melanism in American and British peppered moths

Parallel evolutionary changes in the incidence of melanism are well documented in widely geographically separated subspecies of the peppered moth (Biston betularia). The British melanic phenotype (f. carbonaria) and the American melanic phenotype (f. swettaria) are indistinguishable in appearance, and previous genetic analysis has established that both are inherited as autosomal dominants. This report demonstrates through hybridizations of the subspecies and Mendelian testcrosses of melanic progeny that carbonaria and swettaria are phenotypes produced by alleles (isoalleles) at a single locus. The possibility of close linkage at two loci remains, but the simpler one-locus model cannot be rejected in the absence of contrary evidence.     

Clinal variation, intraspecific heterochrony, and microevolution in the Permian bryozoan Tabulipora carbonaria

Clinal geographic variation across an onshore-to-offshore environmental gradient occurs in the Lower Permian trepostome bryozoan Tabulipora carbonaria collected from three widespread calcareous shales of the Wreford Megacyclothem of Kansas. Separate multivariate statistical tests dclineate significant differences between populations within each shale unit, and between onshore, intermediate, and offshore populations pooled across all three shales, suggesting that even weakly-developed gradients may initiate different intraspecific morphological responses. Growth trajectories for populations along the cline also differ significantly, indicating astogenetic (developmental) heterochrony. Populations from onshore habitats are generally paedomorphic relative to those in more offshore settings, and exhibit pre- and postdisplacement and hypermorphosis in zooecial and acanthostyle characteristics. These heterochronic processes may have increased colonial reproductive potential and the efficiency of water flow for feeding and waste disposal in colonies from onshore habitats. Variation from tightly constrained development (astogenetic plasticity) decreased monotonically in an onshore-to-offshore direction; canalized growth may characterize colonies from more stable offshore habitats, whereas greater flexibility during the growth of colonies from unstable onshore biotopes may have increased their rate of survival. Populations of colonies from stratigraphically successive calcareous shales of the Wreford display patterns of growth that are nearly identical to those found in an offshore-to-onshore direction along the cline. Both clinal and temporal patterns probably resulted from selection for more paedomorphic morphologies in onshore, perhaps unstable, habitats and represent microevolution in T. carbonaria. These local adjustments to environmental conditions may produce variation that affects the rate of macroevolutionary change. □Bryozoa, clines, heterochrony, microevolution, Permian, variability.