Breeding the common bean (Phaseolus vulgaris L.) for resistance to bean common mosaic virus: alternatives to backcrossing

A series of field experiments was undertaken in order to determine whether resistance to bean common mosaic virus (BCMV) could be incorporated into genotypes of the common bean (Phaseolus vulgaris L.) suitable for cultivation in Zimbabwe without recourse to backcrossing. Six inbred genotypes carrying the resistance-conferring alleles at the loci I and Bc-3 were crossed with five locally-adapted inbred genotypes. The first experiment comprised F₃ progeny rows, each derived from a single unselected F₂ plant, the second, F₃ bulks selected for resistance, and the third, a comparison of selected and unselected F₂-derived F₄ lines. The number of days to flowering and to maturity, the incidence of mosaic and necrosis symptoms, seed yield and seed size were recorded. There was evidence that late flowering and maturity were associated with BCMV resistance in some crosses, though not strongly enough to present an obstacle to plant breeding. The incidence of virus symptoms and seed yield were influenced by genetic factors additional to the major resistance genes, and variation in seed yield was present not only between bulk populations of crosses, but also between single-row plots of lines within crosses. This indicates that early-generation selection for yield in the presence of BCMV, even among progeny selected for BCMV-resistace, is likely to be effective. However, the variation in yield among F₄ lines was least in the highest-yielding crosses, which may represent a limit to successful selection for yield. Seed size was partly under additive genetic control, but there was also evidence of non-allelic interactions. There was no association between large seed size, preferred by consumers, and susceptibility to BCMV in the progeny, indicating that the association between these characters in the parent lines is fortuitous and will not present an obstacle to plant breeding. It is noted that a considerable amount of useful genetic information can be obtained without recourse to elaborate crossing schemes, provided that unselected progeny are included in experiments as controls. The evidence presented indicates that resistance to BCMV can be combined with appropriate values of maturity date, yield and seed size without the need for backcrossing.     

Development of common bean (Phaseolus vulgaris L.) lines resistant to the bean pod weevil,Apion godmani Wagner,in Central America

Summary The larva of the bean pod weevil (BPW), Apion godmani Wagner (Coleoptera: Curculionidae), causes serious yield losses in common bean (Phaseolus vulgaris L.) in Mexico and Central America, by consuming the seed as it develops in the immature pod. Resistance to the BPW was identified in bean germplasm of highland Mexican origin, and these sources of resistance were incorporated into a pedigree breeding program to recover locally adapted lines resistant to Bean Common Mosaic Virus and BPW, with commercial grain for Guatemala, Honduras and El Salvador. These lines yielded as well as or better than local cultivars in the absence of the insect, and better than local cultivars when the BPW was present. Resistance appeared to be governed by several genes, and was stable across geographic areas, seasons and planting systems.     

Sources and inheritance of earliness in tropically adapted indeterminate common bean

Summary Nineteen early maturing, indeterminate common bean parents were crossed with a tester line of normal maturity. All parents, F₁ hybrids, F₂'s, and backcross generations were evaluated at CIAT-Palmira, Colombia. All 19F₁ populations flowered and matured almost as early as the respective early parent (mean of 61 vs 59 days). The F₂ populations were slightly less early (63 days), but were still earlier than the mean of the two parents (65 days). Additive effects for days to maturity were usually larger than dominance or interaction effects. Narrow sense heritability for days to flower and maturity were high (greater than 0.67), while heritabilities of absolute and relative duration of pod filling were low (0.17 to 0.50). Also, our data indicated that for each day of increase in earliness, yield potential will decrease 74 kg/ha.     

Linkage of dominant hypersensitive resistance to bean common mosaic virus to seed color in Phaseolus vulgaris L.

Summary Evaluation of Phaseolus vulgaris germplasm bank materials and progenies from a large number of crosses using red- or yellow-colored, BCMV-susceptible bean lines, crossed to purple- or grey/brown-colored, hypersensitive-resistant lines, suggested strong trait association between seed color and BCMV resistance. The cross of red-mottled I⁺I⁺ (susceptible) BAT 1255R to isogenic purple-mottled II (resistant) BAT 1255M was made to study the segregation of the two characters and to recover red-mottled resistant progenies. No recombinant genotypes were observed among 353 F₃ families inoculated with BCMV-NL3, suggesting that linkage of purple-mottled seed color and dominant BCMV resistance is very close.Contribution of the Centro Internacional de Agricultural Tropical.     

Inheritance of temperature sensitivity of the photoperiod response in common bean (Phaseolus vulgaris L.)

Summary Photoperiod response of flowering in common bean (Phaseolus vulgaris L.) is thought to be controlled by the genes Ppd and Hr. However, cultivars also vary in the degree that cooler temperatures reduces their sensitivity to photoperiod. To examine the inheritance of this temperature sensitivity, crosses of cvs. Gordo x de Celaya and Flor de Mayo × Rojo 70 were evaluated at two sites differing in mean temperature and using 12.5-h natural photoperiod or 18-h artificially extended photoperiod. Under 18-h photoperiod at the warmer site, Palmira, no plants of the parents or of the F₂ populations flowered, confirming that the parents were sensitive to photoperiod. Under 12.5-h photoperiod at the cooler site, Popayan, the parents for each cross flowered at similar dates and no segregation for days to flower was observed. However, under 18-h photoperiod, de Celaya and Rojo 70 and the F₁ populations did not flower within 100 days after planting, while the F₂ and F₃ populations showed segregation that was consistent with single gene inheritance, late flowering being dominant. Late flowering at Popayan under 18-h photoperiod indicates a lack of temperature sensitivity, so temperature insensitivity of the photoperiod response was dominant to sensitivity. The name Tip, for temperature insensitivity of photoperiod response, is proposed for this gene, with the recessive form of this gene conditioning earlier flowering at cooler temperatures with long daylengths. It is recognized that the observed segregation patterns could represent the effect of multiple alleles at the Ppd or Hr loci, and studies are proposed to test this possibility with molecular markers and recombinant inbred lines.     

Inheritance of anthracnose resistance in the common bean cultivar Widusa

Snap bean (Phaseolus vulgaris L.) cultivar, Widusa, was crossed to Michigan Dark Red Kidney (MDRK), Michelite, BAT 93, Mexico 222, Cornell 49–242, and TO cultivars to study the inheritance of resistance to anthracnose in Widusa. The segregation patterns observed in six F₂ populations supported an expected 3R:1S ratio suggesting that Widusa carries a single dominant gene conditioning resistance to races 7, 65, 73, and 453 of Colletotrichum lindemuthianum, the causal organism of bean anthracnose. Allelism tests conducted with F₂ populations derived from crosses between Widusa and Cornell 49–242 (Co-2), Mexico 222 (Co-3), TO (Co-4), TU (Co-5), AB 136 (Co-6), BAT 93 (Co-9), and Ouro Negro (Co-10), inoculated with races 7, 9, 65 and 73, showed a segregation ratio of 15R:1S. These results suggest that the anthracnose resistance gene in Widusa is independent from the Co-2, Co-3, Co-4,Co-5, Co-6, Co-9, and Co-10 genes. A lack of segregation was observed among 200 F₂ individuals from the cross Widusa/MDRK, and among 138 F₂ individuals from the cross Widusa/Kaboon inoculated with race 65, suggesting that Widusa carries an allele at the Co-1 locus. We propose that the anthracnose resistance allele in Widusa be named Co-1 ⁵ as Widusa exhibits a unique reaction to race 89 compared to other alleles at the Co-1 locus. RAPD marker A18₁₅₀₀ co-segregated in repulsion-phase linkage with the Co-1 ⁵ gene at a distance of 1.2 cM and will provide bean breeders with a ready tool to enhance the use of the Co-1 ⁵ gene in future bean cultivars.     

Inheritance of resistance to Xanthomonas campestris pv phaseoli in tetary bean (Phaseolus acutifolius)

Summary The F1, F2 and F3 from two crosses within Phaseolus acutifolius were exposed to Xanthomonas campestris pv phaseoli to analyse the inheritance of resistance. The resistant parent, PI 319.443, gave a hypersensitive reaction in leaves and pods with small necrotic lesions. Based on the resistance of F1, the segregation in F2 and the reaction of F3 plants and lines, it is concluded that resistance in leaves and pods is governed by one dominant gene. Comparisons are made with the resistance to X. campetris in P. vulgaris.     

Introgressing resistance to bacterial and viral diseases from the middle American to Andean common bean

The genetic base of cultivars within market classes of common bean (Phaseolus vulgaris L.) is narrow. Moreover, small- and medium-seeded Middle American cultivars often possess higher yield and resistance to abiotic and biotic stresses than their large-seeded Andean counterparts. Thus, for broadening the genetic base and breeding for higher yielding multiple stress resistant Andean cultivars use of inter-gene pool populations is essential. Our objective was to determine the feasibility of introgressing resistance to Been common mosaic virus (BCMV, a potyvirus), and the common [caused by Xanthomonas campestris pv. phaseoli (Xcp) and X. campestris pv. phaseoli var. fuscans (Xcpf)] and halo [caused by Pseudomonas syringae pv. phaseolicola (Psp)] bacterial blights from the Middle American to Andean bean, using gamete selection. Also, we investigated the relative importance of the use of a landrace cultivar versus elite breeding line as the last parent making maximum genetic contribution in multiple-parent inter-gene pool crosses for breeding for resistance to diseases. Two multiple-parent crosses, namely ZARA I = Wilkinson 2 /// ‘ICA Tundama’ / ‘Edmund’ // VAX 3 / PVA 773 and ZARA II = ‘Moradillo’ /// ICA Tundama / Edmund // VAX 3 / PVA 773 were made. From the F₁ to F₅ single plant selection was practiced for resistance to the common and halo bacterial blights in both populations at Valladolid, Spain. The parents and F₅-derived F₆ breeding lines were evaluated separately for BCMV, and common and halo bacterial blights in the greenhouse at Filer and Kimberly, Idaho in 2001. They were also evaluated for the two bacterial blights, growth habit, seed color and 100-seed weight at Valladolid in 2002. All 20 F₁ plants of ZARA I were resistant or intermediate to common and halo bacterial blights in the greenhouse, but their F₂ and subsequent families segregated for both bacterial blights. Segregation for resistant, intermediate, and susceptible plants for common bacterial blight occurred in the F₁ of ZARA II. Simple correlation coefficient for common bacterial blight between the F₁ and F₁-derived F₂ families was positive (r = 0.54 P < 0.05) for ZARA II. From the F₂ to F₅ the number of families resistant to both bacterial blights decreased in both populations. Only four of 20 F₁ plants in ZARA I resulted in seven F₆ breeding lines, and only one of 32 F₁ plants in ZARA II resulted in one F₆ breeding line resistant to the three diseases. None of the selected breeding lines had seed size as large as the largest Andean parent. The use of elite breeding line or cultivar as the last parent making maximum genetic contribution to the multiple-parent inter-gene pool crosses, relatively large population size in the F₁, and simultaneous selection for plant type, seed traits as well as resistance to diseases would be crucial for introgression and pyramiding of favorable alleles and quantitative trait loci (QTL) of interest between the Andean and Middle American beans.     

Inheritance of pod cross-section in beans (Phaseolus vulgaris L.)

Summary OSU 5062, a green bean line with a tendency for pods to become creasebacked (wider than deep) in cross-section, was crossed to near-round-podded cultivars Oregon 83 and Slenderette, oval-podded Bountiful, and flat-podded Roma, to further clarify the inheritance of pod cross-section. Differences among generations in pod cross-section index (PCS), calculated as pod width/depth, were small in the near-round x creaseback crosses (1.07 and 1.01 for Oregon 83 and Slenderette versus 1.20 for OSU 5062). Bountiful (0.63) and Roma (0.50) differed to a greater extent from 5062. Generation means of individual plant PCS of the F₁ and F₂ were almost identical, were almost exactly intermediate between parents, and strongly supported additive inheritance. Backcross data generally supported additive inheritance, but in some cases indicated degrees of dominance of higher PCS. A generation means analysis indicated additive gene action, but deviations from an additive-dominance model were significant in each cross.     

Rooting of bean leaves and use in germplasm evaluation for common bacterial blight resistance

Summary A simple protocol for leaf rooting in beans (Phaseolus vulgaris L.) was developed and used to investigate the reaction of Xanthomonas campestris pv. phaseoli (Xcp), causal pathogen of common bacterial blight disease, in detached versus attached bean leaves. Trifoliate leaves of different sizes (one-third, two-thirds, and fully expanded), either with or without the pulvinus attached to the petioles, were excised from 20 day-old plants of six bean cultivars/lines. Leaf cuttings were cultured in potting medium and then incubated for 5 to 10 days under transparent polyethylene plastic cover in the greenhouse. Roots were readily initiated along the petioles of the leaf cuttings, whether the pulvinus was present or absent. All leaves which were two-thirds expanded and fully expanded developed roots 5 to 7 days after culture. Eighty to 90 percent of the leaves which were one-third expanded formed roots 8 to 10 days after incubation. Laminae of the rooted leaf cuttings were viable and green during the 2 to 3 months period in culture after removing the plastic cover. The common bacterial blight reactions were similar for inoculated attached leaves, detached rooted leaves (inoculated either after or prior to rooting), and moistened detached leaves incubated without rooting. The latter were only usable for evaluation of the Xcp reaction in growth chamber experiments but not under greenhouse conditions. The rooted leaves would be useful for screening bean lines for multiple disease resistance, especially if the pathogens require different environments for disease expression.Abbreviations CBB Common Bacterial Blight - Xcp Xanthomonas campestris pv. phaseoli (Smith) Dye     

Inheritance of resistance to angular leaf spot (Isariopsis griseola Sacc.) in french bean (Phaseolus vulgaris L.)

Summary Angular leaf spot (Isariopsis griseola Sacc.) is a serious disease of French bean in the hills of India and 40 to 70 per cent of the green pods are damaged and rendered unmarketable. Crosses were made between PLB 257, (Phaseolus coccineus L.), a red flowering pole tope, resistant to angular leaf spot, and Contender (Phaseolus vulgaris L.), a highly susceptible commercial cultivar. Studies of the F₁, F₂, and F₃ progenies indicated that PLB 257, carries a recessive gene imparting resistance to angular leaf spot.     

Evaluation of resistance sources and inheritance of resistance in kidney bean to Indian virulences of Colletotrichum lindemuthianum

Summary Forty nine common bean lines comprising of exotic accessions and locally grown cultivars evaluated against Colletotrichum lindemuthianum exhibited differential resistance to its races in Himachal Pradesh, a north-western Himalayan state of India. Some exotic accessions like G 2333, Cornell 49242, PI 207262, Mexique 222, TO, Perry Marrow, Kaboon and Widusa were resistant to more than five Indian races, whereas two Indian accessions KRC-5 and Hans showed resistance to six and four races, respectively. However, nine accessions KRC-8, KR-40, KR-43, KR-81, KR-62-2, KR-90, KR-142, KR-148, and KR-216 were resistant to three races. Race specific resistance has been observed in different bean cultivars. Studies on inheritance of resistance in exotic accession G 2333 and Indian accession, KRC-5 showed that two independent dominant genes conferred resistance in G 2333 to race 3 and 515 and a single dominant gene controlled resistance in KRC-5 to race 775, indicating resistance from these sources is easily transferable to the locally adapted susceptible cultivars.     

Alternative methods of interpreting measurements of resistance to the leafhopper Empoasca kraemeri Ross and Moore in the common bean,Phaseolus vulgaris L.

Summary Appropriate definitions of, and methods of measuring, resistance are sought using Phaseolus vulgaris, a major source of protein in Latin America, and its most important pest, Empoasca kraemeri. The concepts of general vigour and narrow-sense resistance are defined: these two components make up broad-sense resistance. The validity of these definitions, which depend on the inclusion of tolerance as a resistance mechanism, is discussed. In order to distinguish the components, 38 genotypes of P. vulgaris were grown with and without insecticidal proctection, in three replications in a split-plot design, with three planting dates. Seed yield per plant, and a visual score of damage symptoms or vigour, were measured. Three methods of estimating narrow sense resistance are considered: 1) the genotype × insecticide interaction effect in a split-plots analysis, 2) the yield of each genotype unprotected (Y_u) divided by its yield protected (Y_p). and 3) the deviation of each genotype from a regression of Y_u on Y_p. The third method overcomes some objections to the first two, and can be applied to damage and vigour scores as well as to yields. The 38 genotypes differed in narrow-sense resistance according to all three methods, and it appears that in practice Methods 2 and 3 were both valid. They also differed in general vigour. Breeders working with many pests and crops have found that low damage and high yield in the presence of the pest are effective selection criteria. The work reported here supports this view.     

Genetics of resistance to bean golden mosaic virus in Phaseolus vulgaris L.

Summary The genetics of resistance to bean golden mosaic virus (BGMV) was studied in an 8×8 complete diallel cross of common bean, Phaseolus vulgaris L. The 28F₁ hybrids, their reciprocals, and eight parents were artificially inoculated with BGMV under glasshouse conditions. Data were recorded and analyzed for foliar yellowing, plant dwarfing, flower abortion, and pod formation, using a randomized complete block design with two replications.General combining ability (GCA) mean squares were highly significant (P<0.01) and larger than values for specific combining ability for all traits. Reciprocal and maternal effects were nonsignificant. Cultivars Royal Red and Alubia Cerrillos possessed significant negative and positive GCA for all traits, respectively. Porrillo Sintetico also had negative GCA for all traits except plant dwarfing. None of the GCA effects were significant for Great Northern 31 and PVA 1111. Positive associations existed among all traits studied.     

Gamete selection for improving physiological resistance to white mold in common bean

White mold (WM), caused by Sclerotinia sclerotiorum (Lib.) de Bary, is a widespread disease of dry and green bean (Phaseolus vulgaris L.) in North America. Gamete selection (GS) was effective to combine and pyramide resistant genes and quantitative trait loci (QTL) for common bacterial blight. Our objective was to determine the effectiveness of GS to introgress physiological resistance to white mold. Two inter-gene-pool double-cross populations were developed. Selection for WM resistance was practiced from F₁ to F₄. Thirteen selected F_1:5 breeding lines of each population and their four parents were evaluated. Two separate inoculations were made on each plant 1 week apart using a cut-stem method. The WM reaction was scored at 16, 23, and 33 days post inoculation (DPI) using a scale from 1 (no disease) to 9 (severely diseased or dead). In F₁, 52% of Pop I (USPT-WM-1/CORN 601//USPT-CBB-1/92BG-7) and 67% of Pop II (Chase/I9365-25//ABL 15/A 195) susceptible plants were discarded. In F_4, only 1.2% of families from Pop I, and 0.9% for Pop II, survived the selection process. An average of 20.5% gain in WM resistance was obtained for both populations in F₄. Four breeding lines of Pop I had significantly (P = 0.05) lower WM score (4.1–4.6) and four were equal (4.7–4.9) to the best WM-resistant parent 92BG-7 (4.9), while ten breeding lines of Pop II were equal (4.5–4.8) to the best WM-resistant parent A 195 (4.6). Thus, GS was effective for improving WM resistance in common bean.     

Comparison of recurrent and congruity backcrossing for interracial hybridization in common bean

Summary Two common bean (Phaseolus vulgaris L.) cultivars were used for recurrent backcrossing (i.e., repeated backcrossing to one of the parents) to both parents independently and for congruity backcrossing (i.e., backcrossing alternately to both parents). One cultivar, ICA Pijao of race Mesoamerica, was late maturing, with erect type II growth habit and small seeds. The other was Pinto UI 114 of race Durango, an early maturing cultivar with prostrate type III growth habit and medium seeds. One to three recurrent backcrosses (RBC) to both parents, and two rounds of congruity backcrosses (CBC), were made. Thirty-two random lines from each method along with two parents were compared in a reps-in-set design at two locations (Popayán and Quilichao) in Colombia in 1992–1993.Differences between lines derived through different methods were found for seed yield, 100-seed weight, days to maturity, growth habit, and seed color pattern. The RBC to ICA Pijao produced lines with relatively higher seed yield, smaller seed size, and delayed maturity. Similarly, the RBC to Pinto UI 114 produced lines with reduced yield, medium seed size, and earlier maturity. All lines from one, two, and three RBC to Pinto UI 114 had type III growth habit. While most lines from the RBC to ICA Pijao had erect type II Growth habit, a few type III lines were also found in all the RBC. The lines derived from the CBC mostly possessed yield, maturity, and seed characteristics between those of the two parents and lines derived from RBC to both parents independently. They also had more variation for growth habit. To maximize recombination between and retention of desirable traits from distantly related parents used in interracial crosses, use of the CBC is suggested.     

Improvement of large-seeded common bean cultivars under sustainable cropping systems in Spain

Approaches are needed to broaden the genetic base and improve earliness and yield potential of large-seeded beans under sustainable cropping systems. The objective of this research was to develop adapted dwarf bean populations having a commercial seed quality and yield suitable for the production in the South of Europe. The original base populations were produced from crosses between genotypes within each Mesoamerica, Nueva Granada and Peru races, and between Peru and Nueva Granada, and Mesoamerica and Nueva Granada races. Visual mass selection for plant performance was practised in the F₂ and F₃ generations. In the F₄ and F₅, single plants were harvested under two cropping systems (sole cropping and intercropping with maize). From F₄, selection was based on precocity, combined with seed yield and seed commercial type. The F_4:7 selected lines from each original population were compared with their parents and five checks at four environments and two cropping systems. Differences among environments, populations, parents and checks were observed for all traits. Under intercropping with maize, there was a 50% reduction in seed yield. Yield of parents and checks belonging to Andean South American races, intraracial (Nueva Granada × Nueva Granada) and interracial (Nueva Granada × Peru) populations, was higher than that of those of Middle American origin. Intraracial crosses within large-seeded Andean South American (Peru race) and Middle American gene pools (Mesoamerica race) did not produce lines yielding more than the highest yielding parent. Only two large-seeded lines selected from crosses between small- and large-seeded gene pools out-yielded the best parent and check cultivar.     

Growth analysis in a reduced leaf mutant of common bean (Phaseolus vulgaris L.)

Summary A comparison between the Royal Red bean variety and a reduced leaf mutant on the basis of growth indices is reported. The mutants has lower leaf area index (LAI) and leaf area duration (LAD) (–43.7%), higher specific leaf weight (SLW) (on the mean +33.3%) and accumulates less dry matter (–7.7%) than the normal genotype. When the net assimilation rate (NAR) is considered, the mutant is superior to the normal (+62.3%). It is concluded that the unit area of the mutant leaves accumulates dry matter more efficiently than that of the corresponding normal variety.     

Genetic mapping of quantitative resistance to race 5 of Pseudomonas syringae pv. phaseolicola in common bean

Halo-blight is an important worldwide bacterial disease of common bean (Phaseolus vulgaris L.) caused by Pseudomonas syringae pv. phaseolicola. Nine races of the pathogen and five race-specific resistance genes have been previously described. However, a quantitative response to this pathogen has also been described. The objective of this study was to identify halo-blight resistance loci linked to molecular markers that could be used in resistance breeding. Chromosomal regions related to race 5 halo-blight resistance were localized on a genetic map of RAPD and AFLP molecular markers and constructed by the analysis of a “Jules” × “Canela” F₂ progeny. “Jules” shows quantitative resistance to halo-blight and “Canela” is a very appreciated but susceptible Spanish bean landrace. Two QTL for resistance to halo-blight were mapped in two linkage groups. There were four large groups, with 14–22 molecular markers each, five with 4–8 markers each, and three with 2 or 3 markers each.     

Breeding for resistance to bean golden mosaic virus in an interracial population of Phaseolus vulgaris L.

Summary Eighty-three F₂-derived F₈ recombinant inbred lines (RILs) randomly selected from a population (TY 8419) of two common bean genotypes (Pinto UI 114 x ICA Pijao), the two parents, and a control cultivar (Topcrop) were screened for their reaction to bean golden mosaic virus (BGMV) under glasshouse conditions. A randomized complete block design with two replications was used. Data were recorded for disease incidence, foliar yellowing, number of pods and seeds, and yield.Significant (P<0.05) differences among the 83 RILs were recorded for all traits. Of these RILs, 11 lines did not develop any symptom, 24 lines had a mean disease incidence of 8%, 28 lines had a disease incidence of 26.6% and developed intermediate mosaic symptoms, and 20 lines were more susceptible (>50% disease incidence and severe yellowing) than either of the parents, which had 25% to 38% disease incidence and moderate symptoms. Heritability for disease incidence was 0.54±0.01, for foliar yellowing 0.36±0.13, for pods plant^-1 0.69±0.01, for seeds plant^-1 0.54±0.09, and for yield plant^-1 0.49±0.01. Gains from selection (at 20% selection pressure) for these traits were 49.5%, 31.6%, 24%, 0%, and 20%, respectively, over the mean value of all RILs. Correlation coefficients between disease incidence and foliar yellowing and among yield and its components were positive. Disease incidence and foliar yellowing showed negative associations with yield and its components.