The Adaptive Value of Chromosomal Inversions and Climatic Change-Studies on the Natural Populations of Drosophila subobscura from the Balkans.

The adaptive value of the Drosophila subobscura chromosomal inversion polymorphism with regard to environmental effects is well-known. However, the specific details of the inversion adaptations to the global warming scenario deserve to be analyzed. Toward this aim, polymorphism and karyotypes were studied in 574 individuals from Petnica (Serbia) in annual samples taken in June for the period 2019-2022. Comparing the results of Petnica (Cfa: humid subtropical climate) with those from Avala (Serbia: Cfb, temperate oceanic climate) and Font Groga (Barcelona, Spain; Csa: hot-summer Mediterranean climate), significant differences were observed for their chromosomal polymorphism. In Petnica, inversions from U and E chromosomes mainly reacted significantly with regard to temperature, humidity, and rainfall. Moreover, the inversion polymorphism from Petnica (2019-2022) was compared with that from 1995. In this period, a significant increase in mean and maximum temperature was observed. However, to properly explain the observed variations of inversions over time, it was necessary to carefully analyze annual seasonal changes and particular heat wave episodes. Interestingly, yearly fluctuations of U chromosome 'warm'-adapted inversions corresponded with opposite changes in 'non-thermal' inversions. Perhaps these types of inversions were not correctly defined with regard to thermal adaptation, or these fluctuations were also due to adaptations to other physical and/or biological variables. Finally, a joint study of chromosomal inversion polymorphism from many Balkan populations of D. subobscura indicated that different climatic regions presented distinct composition, including thermal-adapted inversions.

Adaptation of Drosophila subobscura chromosomal inversions to climatic variables: the Balkan natural population of Avala.

The adaptive value of chromosomal inversions continues raising relevant questions in evolutionary biology. In many species of the Drosophila genus, different inversions have been recognized to be related to thermal adaptation, but it is necessary to determine to which specific climatic variables the inversions are adaptive. With this aim, the behavior of thermal adapted inversions of Drosophila subobscura regarding climatic variables was studied in the natural population of Avala (Serbia) during the 2014-2017 period. The results obtained were compared with those previously reported in the Font Groga (Barcelona, Spain) population, which presents different climatic and environmental conditions. In both populations, it was observed that most thermal adapted inversions were significantly associated with the first, second or both principal components, which were related with maximum, minimum and mean temperatures. Moreover, a significant increase over years (2004-2017) for the minimum temperature was detected. In parallel, a significant variation over time in Avala was only observed for the frequencies of 'warm' and 'non-thermal' adapted inversions of the U chromosome. However, stability in the chromosomal inversion polymorphism was observed for the 2014-2017 period which might result from the temporal span of the study and/or selective process acting on the population.

Rate of change for the thermal adapted inversions in Drosophila subobscura.

The changes of chromosomal inversion polymorphism composition of Drosophila subobscura in samples from Apatin (Serbia) were studied in a 24-years interval (1994-2018). The variation was significant for all autosomes and directional, increasing the inversions considered as 'warm', whereas those reported as 'cold' decreased. Furthermore, the Chromosomal Thermal Index (CTI), which allows studying the thermal adaptation of the whole karyotype increased significantly in that period of time. These results were in agreement with the indicators of global warming in Apatin: a trend to increase of the mean, maximum and minimum (this latter even significant) temperatures, and an erratic pattern of rainfall (also usual in global warming). The deviations from the Wright-Fisher model of genetic drift were used to consider the possible effect of migration or selection as evolutionary factors responsible for the change in inversion frequencies. To quantify approximately the rate of change in the frequencies, for each kind of inversions ('cold', 'warm' and 'non-thermal adapted'), the difference in frequency between the Apatin samples obtained in 1994 and 2018 was computed and then it was divided by the number of years elapsed. This rate was always higher (from twice as many as thirty times more depending on the autosome) for thermal adapted inversions ('cold' or 'warm') than the 'non-thermal' adapted. From this study, it could be concluded that the chromosomal inversions of D. subobscura could change (in composition and frequencies) in a predictable direction and a rather 'rapid' rhythm to adapt to the global warming scenario.

Chromosomal Thermal Index: a comprehensive way to integrate the thermal adaptation of Drosophila subobscura whole karyotype.

Drosophila has demonstrated to be an excellent model to study the adaptation of organisms to global warming, with inversion chromosomal polymorphism having a key role in this adaptation. Here, we introduce a new index (Chromosomal Thermal Index or CTI) to quantify the thermal adaptation of a population according to its composition of "warm" and "cold" adapted inversions. This index is intuitive, has good statistical properties, and can be used to hypothesis on the effect of global warming on natural populations. We show the usefulness of CTI using data from European populations of D. subobscura, sampled in different years. Out of 15 comparisons over time, nine showed significant increase of CTI, in accordance with global warming expectations. Although large regions of the genome outside inversions contain thermal adaptation genes, our results show that the total amount of warm or cold inversions in populations seems to be directly involved in thermal adaptation, whereas the interactions between the inversions content of homologous and non-homologous chromosomes are not relevant.

Inbreeding and thermal adaptation in Drosophila subobscura.

Using a well-adapted Drosophila subobscura population (Avala, Serbia), a drastic experiment of inbreeding was carried out to assess whether the expected level of homozygosity could be reached or if other evolutionary forces affected the process. In general, no significant changes of inversion (or arrangement) frequencies were detected after 12 brother-sister mating generations. Furthermore, no significant differences were obtained between observed and expected (under the inbreeding model) karyotypic frequencies. Thus, these results seemed to indicate that the main evolutionary factor in the experiment was inbreeding. However, in the G12 generation, complete chromosomal fixation was reached only in two out of the eight final inbred lines. In these lines, the chromosomal compositions were difficult to interpret, but they could be likely a consequence of adaptation to particular laboratory conditions (constant 18 °C, food, light period, etc.). Finally, in a second experiment, the inbred lines presented higher fertility at 18 °C than at 13 °C. Also, there was a significant line effect on fertility: inbred line number 6 (A1, J1, U1+2; U1+2+6, E8, and O3+4+7) presented the highest values, which maybe the result of an adaptation to laboratory conditions. Thus, the results obtained in our experiments reflect the adaptive potential of D. subobscura inversions.

Changes in chromosomal polymorphism and global warming: The case of Drosophila subobscura from Apatin (Serbia).

In this study, chromosomal inversion polymorphism data for a natural population of Drosophila subobscura from a swampy region near the town of Apatin (Serbia) were compared with data for the same population collected approximately 15 years earlier. The pattern of chromosomal inversion polymorphism changed over time. There were significant increases in the frequency of characteristic southern latitude ("warm" adapted) chromosomal arrangements and significant decreases in the frequency of characteristic northern latitude ("cold" adapted) chromosomal arrangements in the O and U chromosomes. The chromosomal arrangements O(3+4) and O(3+4) (+) (22) (derived from the O(3+4) arrangement) showed significant increases in 2008 and 2009 with regard to the 1994 sample. There was also a significant increase (∼50%) in the U(1) (+) (2) arrangement, while U(1+8) (+) (2) (a typical southern arrangement) was detected for the first time. Since the Apatin swampy population of D. subobscura has existed for a long time in a stable habitat with high humidity that has not been changed by man our results indicate that natural selection has produced chromosomal changes in response to the increase in temperature that has occurred in the Balkan Peninsula of central southeastern European.

Changes in chromosomal polymorphism and global warming: the case of Drosophila subobscura from Apatin (Serbia)

In this study, chromosomal inversion polymorphism data for a natural population of Drosophila subobscura from a swampy region near the town of Apatin (Serbia) were compared with data for the same population collected approximately 15 years earlier. The pattern of chromosomal inversion polymorphism changed over time. There were significant increases in the frequency of characteristic southern latitude ("warm" adapted) chromosomal arrangements and significant decreases in the frequency of characteristic northern latitude ("cold" adapted) chromosomal arrangements in the O and U chromosomes. The chromosomal arrangements O3+4 and O3+4+22 (derived from the O3+4 arrangement) showed significant increases in 2008 and 2009 with regard to the 1994 sample. There was also a significant increase (~50 percent) in the U1+2 arrangement, while U1+8+2 (a typical southern arrangement) was detected for the first time. Since the Apatin swampy population of D. subobscura has existed for a long time in a stable habitat with high humidity that has not been changed by man our results indicate that natural selection has produced chromosomal changes in response to the increase in temperature that has occurred in the Balkan Peninsula of central southeastern European.

Viabilities of Drosophila subobscura homo- and heterokaryotypes at optimal and stress temperatures. I. analysis over several years.

The interactions of lethal and non-lethal genes and their contributions to the viability of Drosophila inversion karyotypes are not well understood. This is especially true under variable environmental conditions. Here we examine the viability of natural chromosomal O-inversion homo- and heterokaryotypes in a D. subobscura population from Avala Mountain, Serbia. The observations we report were performed at a range of temperatures over several years. The heterotic effect of O-lethal heterozygotes on viability was found to be independent of the effects of inversion backgrounds and temperature. Positive epistatic interactions of lethal, mildly deleterious (subvital) and quasinormal (normal) genes were found in O-inversions in heterokaryotypes but not in homokaryotypes. These interactions were independent of temperature. This finding could explain the limitation of the genetic load in D. subobscura populations. In the population analyzed, annual fluctuations in the frequencies of certain chromosomal arrangements, karyotypes and non-lethal chromosomes under cold-stress temperatures seemed to indicate a correlation between these polymorphisms and environmental conditions. Our results indicate that there is a response in tolerance to extreme temperatures that may be due to natural selection. The differences in mean viability between some O-inversion karyotypic combinations indicate that there are differences in their tolerance to variable temperatures. All our results suggest that both frequency-dependent and supergene selection are mechanisms that protect O-chromosomal inversions. Chromosomal inversions may be genetically differentiated and coadapted complexes in D. subobscura populations.

Viabilities of D. subobscura homo- and heterokaryotypes at optimal and stress temperatures. II. Seasonal component analysis.

Here we report the viability of natural chromosomal O-inversion structural homo- and heterokaryotypes in a population of D. subobscura from Avala Mountain, Serbia, subjected to cold and heat stress during spring 2004. These results were compared with those observed in the same population in September 2004. We detected protection of a great number of lethal genes in O-chromosome lethal heterozygous combinations, independently of inversion backgrounds and temperature stress. The positive epistatic interactions of mildly-deleterious genes observed only in O-inversion heterokaryotypes (at distinct temperatures) and lethal genes in O-inversion homokaryotypes (under temperature stress only) could be responsible for reducing the genetic load. The seasonal fluctuations in frequencies of several chromosomal arrangements and karyotypes indicate a seasonal adaptation to changeable environmental conditions. The sample of D. subobscura collected in spring 2004 was tolerant to cold but not to heat, and these results point to tolerance to extreme low temperatures, possibly as a result of natural selection. Also, a non-random distribution of distinct combinations of viability classes among several O-inversion karyotypes (under both optimal conditions and heat stress) was observed. This observation could be taken as evidence of co-adaptation: different capacities of several O-inversion karyotypes with different combinations of genes to tolerate a range of temperatures. We conclude that frequency-dependent and supergene selection are balancing selection mechanisms and are responsible for the protection of chromosomal inversions in natural populations of D. subobscura.

The genetic structure of Balkan populations of Drosophila subobscura.

Although Drosophila subobscura has been a model organism for European and American population geneticists, little information is available on the genetic structure of its natural populations. In this paper we report the estimates of some population parameters. We have used data from lethal allelism tests in four Balkan populations (Kamariste, Djerdap and Petnica in Serbia and Zanjic in Montenegro). In all populations, lethal genes were found to have a deleterious effect on heterozygotes. The N(e) values varied greatly from 370 (Petnica) to 19413 (Kamariste), depending on the habitat conditions and some environmental factors. Finally, gene flow between the Balkan populations was detected by the estimates of N(m) (from 4.68 in Petnica to 106.2 in Kamariste) and m (from 0.0041 in Djerdap to 0.0126 in Petnica). These results agree with those obtained in a previous study where the frequencies of allelism between populations were greater than predicted by independently arising lethal genes.

Viabilities of originally natural O-chromosomal inversion homo- and heterokaryotypes in Drosophila subobscura.

The influence of epistatic interactions of lethal and non-lethal genes upon viability of Drosophila inversion karyotypes is poorly known. In this paper we present comparative results for viabilities of 21 originally natural O-inversion homo- and 38 heterokaryotypes in a D. subobscura population. We observed strong heterotic effect in viability of O-lethal heterozygotes irrespective of different inversion backgrounds, which indicates a mechanism for protection of a great number of lethal genes. In O-non-lethal heterozygotes the heterotic effect in viability was absent irrespective of different inversion backgrounds. Our results showed a great extent of genetic load and high abundance of O-chromosomal arrangements in the population analyzed. It belongs to the set of central European populations. An epistasis of lethal genes present in O-inversion hetero- and not present in O-inversion homokaryotypes of moderate or low frequencies could be good example for co-adaptation of chromosomal inversions with regard to the genetic load. This represents a more efficient mechanism for limitation of genetic load than alternative mechanisms for protection of lethals. Except for lethal genes, possible epistatic interactions of mildly deleterious (subvital) genes, could also be responsible for limiting the extent of genetic load in natural D. subobscura populations. We demonstrated a non-random distribution of several combinations of viability classes among different O-inversion homo- and heterokaryotypes. As a consequence of that, the viabilities of the O-inversion homokaryotypes compared to heterokaryotypes were significantly higher at low frequencies than in moderate or high frequencies. This shows frequency-dependence as a mechanism of balancing selection for protection of O-chromosomal inversions in natural D. subobscura populations. In addition, the viabilities of the O-inversion homokaryotypes of lower frequency, compared to homokaryotypes of moderate or high frequency, were significantly higher. This again indicates the existence of supergene selection as another mechanism for protection of chromosomal inversions, as co-adapted complexes in natural D. subobscura populations.