Genetic load affecting chromosomes II and III in two natural populations of Drosophila melanogaster from Mexico.

Salceda, Victor M.¹, and Aluisio J. Gallo² .  ¹ Departamento de Biología, Instituto Nacional de Investigaciones Nucleares. Apartado Postal 18-1027. Mexico, 11801, D.F. MEXICO; E-mail: vmss@nuclear.inin.mx.; ² Universidade Estadual Paulista "Julio de Mesquita Filhio", Departamento de Biología. Sao José do Río Preto, Sao Paulo, BRASIL.

     A great many studies on the genetic load in natural populations of Drosophila have been carried out by many authors with reference to the frequency of lethal and semilethal genes found in one of the chromosomes in the genome. There exist fewer studies in which two or more chromosomes are analyzed at the same time (e.g., Pavan et al.,1951;  Dobzhansky and Spassky, 1953; Spiess and Allen, 1961; Dobzhansky et al., 1963; Band and Ives, 1963; Spassky, Dobzhansky and Anderson, 1965; Hoenigsberg and Navas, 1965; Wallace, Zouros and Krimbas, 1966; Paik, 1966; and Allen, 1966, 1969). As a group these authors found information on natural populations of  D. melanogaster, D. persimilis, D. pseudoobscura and D. willistoni, and the information has been reported by them in reference to the amount of the load found in different species and populations.  This load could be restricted to a single chromosome or in some cases to two of them. In special situations they deal with particular phenomena, such as epistatic effects (Spassky, Dobzhansky and Anderson, 1965) or to the measurement of the variability (Dobzhansky and Spassky, 1951; Spiess and Allen, 1961), or other aspect of interest for the author.

     Among the studies dealing with the load due to interaction between two or more chromosomes, we have those of Wallace, Zouros and Krimbas (1966) and Paik (1966) as representatives of this kind of analysis.

     All of the above studies were done outside of Mexico. Those investigations dealt principally with an analysis of frequencies of deleterious chromosomes in different localities and at different seasons.  In the present case, we are dealing with the effect of genetic load in two natural populations of D. melanogaster from Mexico.

Material and Methods

     Samples of D. melanogaster from two localities (Coyoacan, D.F., Mexico City, with an altitude above sea level of 2250 m, and Veracruz City in the State of Veracruz located at sea level) were analyzed using the Cy L / Pm ; Ubx / Sb technique.

     Groups of 20 males were studied weekly. Each male was crossed with three virgin females of the marker stock SM1 al² Cy sp² /  Pm, In(3RL) ; Ubx¹³⁰, Ubx¹³⁰ e^s / Sb. A single F1 Cy L / + ; Sb / + male was back-crossed with three virgin females from the marker stock. In the F2, five pairs of flies of the Cy L / + ; Sb / + constitution were mated inter se. The flies were allowed to oviposit during four days, transfered to a second bottle for an additional four day period and then discarded. Starting 12 days after the initiation of the culture, emerged flies were counted every other day until no more appeared. It is expected that, in the absence of a significant genetic load, four kinds of F3 flies will have been produced and in the proportions:  4 Cy L / + ; Sb / + : 2 Cy L / + ; + / + : 2 +/+ ; Sb / + : 1 +/+ ; +/ +. All the cultures were kept at 25 ± 1°C, using as a food the regular corn meal - sugar - agar culture medium employed at the laboratory.

     In the case of the population from Coyoacan located at a distance of about 5 km from the laboratory, males were collected each week prior to the crosses being done. Since Veracruz is approximately 400 km from the laboratory, a four-day collection of 314 males and 264 females was made. The flies were then introduced into a population cage and allowed to increase in number for two generations. Then 20 males were extracted for analysis every week. A minimum of 200 males were tested from each population. Analysis of 135 genomes from Coyoacan and 148 from Veracruz, were carried out.

Table 1.  Classification of the different categories of chromosomes. Category of Chromosomes Chromosomes II and III, independently, % of expected wild-type flies Interaction of chromosomes II and III, % of expected wild-type flies Lethal 0 0 Extreme semilethal 1-3 1-2 Semi-lethal 4-17 3-8 Sub-vital 18-25 9-16 Normal 26-40 17-23 Super normal >40 >23 Table 2.  General behavior in the viability of the populations.  Observed and expected numbers of flies according to the mendelian proportions of the experimental crosses. Population Cy L /+;Sb/ + 4 + /+ ; Sb / + 2 Cy L / + ; +/+ 2 + / + ; + / + 1 Coyoacan Observed 22012 8549 7213 2527 Expected  17911 8955 8955 4477 Veracruz Observed 23609 6982 7429 2093 Expected 17828 8914 8914 4457 * all comparisons are different at P<0.0001

Results

     When all the counts were completed, we assigned chromosomes to different classes according to their relative viabilities.  The classification system is shown in Table 1.

     Differences in classification (column 2 vs. column 3) are due to the fact that when a chromosome is analyzed independently for its viability, the expected ratio with respect to the control  Cy L / + ; Sb / + heterozygotes is 2 :1, and when interactions are measured this proportion is 4:1.

     Table 2 shows the corresponding data when the total number of flies is summarized according to genotype and a corresponding Mendelian proportion. This allows us to evaluate the overall viability of the populations from the two different  areas.

     Table 3 shows the corresponding data when the total number of flies is summarized according to genotype and a corresponding Mendelian proportion. This allows us to evaluate the overall viability of the populations from the two different  areas.

     The numbers and frequencies of chromosomes II and III carrying deleter-ious genes and II-III interactions found in each population are shown in Table 3.

    The actual number of lethals found per chromo-some and their interaction for each population studied is shown in Table 4.

Discussion

Analysis of Table 2 indicates that the populations tend to carry appreciable loads of genetic variability, since virtually all observed values are higher than the expected, in terms of the number of flies counted in each category.

    Summing the lethals, extreme semi-lethals (considered by some people as lethals; Wallace, 1970), and semilethals for each chromosome and their interactions, we note that the value for the population from Coyoacan concerning chromosome II is 25.91 as shown in Table 3, which is in the range of those found by the majority of the authors that have studied this chromosome (i.e., 12 to 30%).

In the case of the population from Veracruz, the corresponding value is 40.52%, quite high according to the values from other studies.  If  semilethals are excluded, the value is 24.31% and in the range of those populations previously studied by other authors, suggesting the possibility that the difference is primarily due to the high frequency of semilethals found in the present population of Veracruz. If the same assumption is made for the population from Coyoacan (semilethals omitted) the value of the genetic load is 10.37% and in the very low end of those reported previously.

Table 3.  Number and frequency of deleterious chromosomes II and III in each population and their interaction. Populations Kind Coyoacan 135 genomesChromosome Veracruz 148 genomesChromosome II III II and III II III II and III Lethals (6) 4.44 (4) 2.96 (27) 20.00 (19) 12.83 (0) 0.0 (36) 24.32 Extreme semilethal (8) 5.92 (4) 2.96 (10) 7.40 (17) 11.48 (0) 0.0 (18) 12.16 Semilethal (21) 15.55 (28) 20.74 (27) 20.00 (24) 16.21 (32) 21.62 (27) 18.24 Subvitals (12) 9.88 (19) 14.07 (37) 27.40 (17) 11.48 (51) 34.45 (42) 28.24 Normals (73) 54.07 (76) 56.29 (29) 21.48 (60) 40.54 (59) 39.86 (19) 12.83 Super normals (15) 11.11 (4) 2.96 (5) 3.70 (11) 7.43 (6) 4.05 (6) 4.05

    When similar analyses are carried out on data from chromosome III, the Coyoacan population yields a value of 26.66% and Veracruz 21.62%. In both cases the corresponding frequencies for lethals and semilethals together are included in the range of values reported in the literature. An interesting finding in the  population from Veracruz (which we cannot explain yet) is the apparent absence of lethals/extreme semilethals in chromosome III (none in 148 chromosomes analyzed as opposed to 36 in 148 chromosome II analyzed).  The statistical significance of it is shown in Table 4.  Here the differences are significant. In contrast, lethals and extreme semilethals were found in both chromosomes II and III in the population from Coyoacan 14/135 = 10.37% in the second chromosome and 8/135 = 5.92% in the third chromosome.  They are also shown in Table 4.

 

Table 4.  Differences for the number of lethas found between the two populationsfor chromosomes II, III and their interaction II-III. Chromosome II Chromosome III Interaction II and III Veracruz n=148 36 0 54 Coyoacan n=135 14 8 37 C2 7.78* 8.77* 1.88 * P<0.001

    Finally results repor-ted by others are compared with those from the present report as shown in Table 5.  Here we must make reference only to those studies done in D. melanogaster (Band, 1963; Hoenigsberg and Navas, 1965; Wallace, Zouros and Krimbas, 1966; and Paik, 1966).  These authors reported for chromosome II values that varied from 15.20% in Korea up to 35.90% in USA.  In Colombia, depending on the population, they ranged from 25.34 to 48.09%, with an average value of 36.71%.  As could be seen, our populations show values that fall into the limits reported by those authors. It is important to mention that the differences in the two populations here reported, with respect to the load for the second chromosome, is significant as shown in Table 4.

    With respect to the third chromosome also, the values found for us are included within the range reported by those authors, since they found values that varied from as low as 18.40% in Korea up to a highest of 40.10% in the USA. For this chromosome the values found for us are quite similar and not significant.

Table 5.  Percentage of deleterious (lethal plus semiletals) reported for chromoses II, III and their interactionin several populatios of Drosophila as reported by different authors. Population II Chrom. Species II Chrom. Interaction Reference Coyoacan 25.9 D. melanogaster 26.7 47.4 This report Veracruz 40.5 D. melanogaster 21.7 54.7 This report USA 35.9 D. melanogaster 40.1 ---- Band, 1963 Colombia 36.7 D. melanogaster ---- ---- Hoenigsberg, 1965 Colombia 26.6 D. melanogaster 26.6 32.8 Wallace, 1966 Korea 15.2 D. melanogaster 18.4 16.8 Paik, 1966

   As for the interaction, the values found by Wallace, Zouros and Krimbas (1966) in Colombia and the one by Paik (1966) in Korea are our points of reference for comparisons. In Colombia the value is 32.80% and in Korea 16.80%.  Both values are quite a bit lower and, with respect to our findings, statistically different.  Our populations also are different among them.

    Of course the biological significance of it could be only ascribed to the very particular background of the populations in which the environment has much influence, and it is reflected in the differences found either by us or by the other authors.

    Acknowledgments: The authors are grateful to Dr. Louis Levine for reviewing the manuscript. A.J. Gallo thanks the Fundaçao de Amparo a Pesquisa do Estado de Sao Paulo, Brasil, for granting him a fellowship to undertake the present study. We acknowledge the support and interest of the Colegio de Postgraduados during the course of our investigation.

    References:  Allen, A.C., 1966, Genetics 54: 1409-1422; Allen, A.C., 1969, Genetics 63: 629-637;  Band, H.T., and P.T. Ives 1963, Can. J. Gent. Cytol. 5: 351-357;  Dobzhansky, Th., and B. Spassky 1953, Genetics 38: 471-484;  Dobzhansky, Th., A.S. Hunter, O. Pavlovsky, B. Spassky, and B. Wallace 1963, Genetics 48: 91-103;  Hoenigsberg, H.F., and Y.G. Navas 1965, Evolution 19: 506-513;  Paik, Y.K., 1966, Japan J. Genet. 41: 325-333;  Pavan, C., A.R. Cordeiro, N. Dobzhansky,  Th. Dobzhansky, C. Malogdowkin, B. Spassky, and M. Wendel 1951, Genetics 36: 13-30;  Spassky, B., Th. Dobzhansky, and W.W. Anderson 1965, Genetics 52: 653-664;  Spiess, E.B., and A.C. Allen 1961, Genetics 46: 1531-1553;  Wallace, B., 1970, Genetic Load. Its Biological and Conceptual Aspects. Prentice Hall, Inc., Englewood Cliffs. N.J.;  Wallace, B., E. Zouros, and C.B. Krimbas 1966, Am. Naturalist 100: 245-251.