Heredity 55 (1985)335—339 The Genetical Society of Great Britain Received 20 March 1985 Genetic variation and selection for fertility in the fungus Cochliobolus heterostrophus James A. Kolmer and Department of Plant Pathology, North Carolina State Kurt J. Leonard University; United States Department of Agriculture, Raleigh N.C. 27695-7616, U.S.A. The response to selection for perithecial number in Cochliobolus heterostrophus, a heterothallic ascomycete, is described. Thirteen MATA isolates and 12 MATa isolates of the fungus were mated in all possible combinations and evaluated for fertility. Progeny from some of these matings segregated for a defective perithecial allele. Selection for increased numbers of mature perithecia was carried out for six generations of intermatings among four to six parents in each generation which were selected from approximately 24 progeny from the preceeding generation. After six generations the numbers of perithecia had increased by nearly a factor of four. The proportion of additive genetic variance decreased linearly in the selection generations. The increase in perithecia was confirmed in concurrent mating tests among subpopulations from each selection generation. INTRODUCTION Selection experiments have been used to demonstrate polygenic variation for various traits Quantitativegenetic variation has received little in fungi. In Aspergillus nidulans, penicillin titer was attention in studies of the genetics of plant doubled after four generations of recurrent selec- pathogenic fungi although it is thought to be tion (Merrick, 1976). Selection in both directions important in determining the parasitic abilities of over several generations for hyphal growth rate these organisms. Cochliobolus heterostrophus was effective in Neurospora crassa (Papa, Srb and Drechsler, the causal agent of southern corn leaf Federer, 1966). Similar results were found for blight, is a heterothallic fungus with two mating growth rate in Schizophyllum commune (Simchen, type alleles, MATA and MA Ta, required for sexual 1966). The response to selection in these studies reproduction in pairings between haploid isolates was evidence of polygenic variation influencing (Nelson, 1957). Nelson (1959a) observed differen- these traits. The present study was undertaken to ces in numbers of perithecia produced in matings determine the nature of the genetic control of of sexually compatible isolates of the fungus, but perithecial number in Cochliobolus heterostrophus did not quantitatively analyse these heritable Drechsler. The extent of polygenic variation pres- differences in fertility. Nelson (1959b) later iden- ent in the fungus which influences fertility was tified a single gene in naturally occurring isolates tested by means of recurrent selection for increased that can eliminate or greatly reduce perithecial perithecial number. production when it is present in both members of a pair of sexually compatible isolates of the fungus. In studies with other species of Cochliobolus, MATERIALS AND METHODS Nelson and others demonstrated that fertility is controlled by genes other than the two alleles at Initial population the mating type locus (Nelson, 1961; 1964; 1970; Thirteen MATA and 12 MATa isolates were Kline and Nelson, 1968), but the role of polygenic chosen from a group of conidial isolates collected variation was not rigorously examined. from diseased corn leaves from fields in North Paper number 9714 of the journal series of the North Carolina Carolina or from first generation ascospore pro- Agricultural Research Service. geny obtained from crosses of such isolates. The 336 J. A. KOLMER AND K. J. LEONARD 25 isolateswere paired in all possible combina- Table 1 North Carolina Design II. Sources of variation and tions. All parental and progeny isolates in this expected mean squares study were stored at 4°C in potato dextrose agar (PDA) slants. The fungal isolates were grown on Source d.f. Expected mean square PDA containing 10 g dextrose per litre for 4 days Isolates MATA A-i V+(4)VA(4a)VMATA after which small (2-4 mm) blocks of agar with Isolates MATa a-i V,+(4)Vfl<U(4A)VMAT the fungus were transferred to petri dishes filled tvJATAxMATa (A-I)(a-1) V.,--(4)VAfl with Sach's agar. Isolates of compatible mating Error Aa(4-1) V types were placed opposite of each other across autoclaved, 1 cm diametre, senescent corn leaf disks in the Sach's agar petri dishes. Four leaf disks as modified for inbred lines (Eberhart, Moll, and per petri dish were used for each MATA x MATa Cockerham, 1966). The expected mean squares are isolate combination. All crosses were kept in the listed in table 1. The combined main effects of dark at 25°C for 14 days. Mature perithecia that isolate MATA and isolate MATa determined the developed on the corn leaf disks were counted, additive genetic variance and the MATA x MATa term defined the additive x additive genetic vari- with each leaf disk representing one replication. ance in the initial population and all selection generations. Selectionfor fertility Thetwo to three most fertile MATA and the two to three most fertile MATa isolates (as expressed RESULTS by the average number of mature perithecia the isolate produced in crosses with all isolates of the Initial population compatible mating type) were selected as parents Matings among the initial population produced an for the next generation. Ascospore progeny were overall mean of 4419 perithecia per leaf disk with isolated from the selected crosses, evaluated for a standard error of 0-59. Genetic variation accoun- mating type, and then crossed in all possible com- ted for 79 per cent of the total variation in the binations to form the next selection generation. initial population. Eight MATA isolates and four Approximately 12 isolates of each mating type MATa isolates in this initial population had a were used in each selection generation. Selection defective allele at the perithecial locus (Leonard, was carried out for six generations. 1978; Nelson, 1959b). Crosses in which both parents had the defective perithecial allele pro- Evaluation duced an average of 1579±068 perithecia per of subpopulations leaf disk; crosses in which one parent had the Toprovide a control for the selection experiment, perithecial allele produced an average 4109 076 subpopulations of each generation were evaluated perithecia per leaf disk; and crosses in which in common environments. Four or five isolates of neither parent had the defective perithecial allele each mating type in each generation were ran- had an average of 722± 173 perithecia per leaf domly chosen and crossed with compatible isolates disk. Thus, the perithecial allele appeared to act of the same generation. In three separate tests, additively in the paired isolates to determine the subpopulations from the initial generation and number of perithecia per leaf disk. The perithecial selection generations one, two, and three; gener- locus accounted for 58 per cent of the genetic ations three, four and five; generations four, five variation in the initial population, and the balance and six, respectively, were evaluated in common of the variation was accounted for by polygenic environments. effects. Additive genetic variance (including the additive effects of the perithecial gene) accounted Analysisof genetic variance for 83 per cent of the genetic variation, with the rest of the genetic variation attributed to additive x Themodel, Number of perithecia per isolate pair = additive genetic variation. meaneffects of isolate MATA+mean effects of isolate MATA+specific interactions between iso- late MATA and isolate MA Ta, was used to esti- Responseto selection mate the additive and additive x additive genetic Theresponse to selection for increased numbers variances in all generations. The genetic variance of perithecia over six generations is illustrated in was partitioned using North Carolina Design II fig. 1. The analysis of variance for each generation SELECTION IN C. HETEROSTROPHUS 337 (I) Table 2 North Carolina Design II. Analysis of variance for a 180 number of perithecia in initial and selection populations U- 160 UJ 140 GenerationSource -J 120 of of Mean selection variation d.f. square 0Lii 0 Isolates MATA 12 14,084* 0 60 11 LU Isolates MAYa 13,026* 40 MATAXMATa 120 614* I-.I 20 M.S.E.t 454 216 0LU 1 Isolates MATA 13 3,840* 1 2 3 4 5 6 /// Isolates MAYa 11 18,835* GENERATIONSOF SELECTION MATAxMATa 131 1,018* Figure 1The response to selection based on additive genetic M.S.E. 457 436 variance over six generations. 2 Isolates MATA 14 7,196* Isolates MATa 8 9775* MATAxMATa 107 1,347* is presented in table 2. The means, selection differ- M.S.E. 356 748 entials, and components of genetic variation are 3 Isolates MAYA 12 5,084* presented in table 3. With six cycles of selection Isolates MAYa 11 13,628* the mean number of perithecia per leaf disk MATAXMATa 127 1,735* increased from 90 to 165 and the standard errors M.S.E. 449 842 ranged from 024 to 326. 4 Isolates MAYA 10 8,031* The selected isolates in the initial population Isolates MAYa 10 20,712* all had the effective perithecial allele, thus fixing MAYAXMAYa 88 3,712* the allele in the subsequent selection generations. M.S.E. 315 1,467 All selection after the first cycle utilised polygenic 5 Isolates MAYA 12 2,465* variation. The proportion of additive genetic vari- Isolates MAYa 10 6,582* MAYA XMAYa 107 1,791* ance relative to total genetic variance in the initial M.S.E. 394 767 and selected populations decreased linearly from 083, to 023 over the six generations (fig. 2). 6 Isolates MAYA 11 13,689* Isolates MAYa 6 21,573* MAYA XMATa 53 7,962* Evaluationof subpopulations in M.S.E. 210 1,033 common environments * Significantat 005. tMeansquare error. Subpopulationsconsisting of isolates randomly sampled from the initial and selected populations were evaluated in three separate tests shown in fig. confirmed in these tests. In a separate test, sub- 3.