Understanding pollen and ovule fertility as factors influencing fruit and seed set is important in cassava breeding. Extended daylength with red light (RL) and plant growth regulators (PGRs) have been used to induce flowering and fruit set in cassava without any reference to effects on pollen viability or ovule fertilizability. This study investigated the effects of field-applied RL and PGR on pollen viability and ovule fertilizability. Panels of cassava genotypes with early or moderate flowering responses were used. RL was administered from dusk to dawn. Two PGRs, 6-benzyl adenine (BA), a cytokinin and silver thiosulphate (STS), an anti-ethylene, were applied. Pollen viability was assessed based on pollen grain diameter, in vitro stainability, in vivo germinability, ovule fertilizability, and ploidy level. Treating flowers with RL increased the pollen diameter from 145.6 in control to 148.5 µm in RL, 78.5 to 93.0% in stainability, and 52.0 to 56.9% in ovule fertilizability in treated female flowers. The fruit set also increased from 51.5 in control to 71.8% in RL-treated female flowers. The seed set followed a similar trend. The ploidy level of pollen from RL-treated flowers increased slightly and was positively correlated with pollen diameter (R2 = 0.09 *), ovule fertilization (R2 = 0.20 *), fruit set (R2 = 0.59 *), and seed set (R2 = 0.60 *). Treating flowers with PGR did not affect pollen diameter but increased stainability from 78.5% in control to 82.1%, ovule fertilizability from 42.9 to 64.9%, and fruit set from 23.2 to 51.9% in PGR-treated female flowers. Combined BA + STS application caused the highest ovule fertilizability, fruit, and seed set efficiency. These results show that RL and PGR treatments increase pollen viability and ovule fertilizability. This is important for planning pollination strategies in cassava breeding programmes.

Understanding pollen and ovule fertility as factors influencing fruit and seed set is important in cassava breeding. Red light (RL) and plant growth regulators (PGRs) have been used to induce or enhance flowering and fruit set in cassava without any reference to effects on pollen viability or ovule fertilizability. This study investigated effects of field-applied RL and PGR on pollen viability and ovule fertilizability. Panels of cassava genotypes, with early or moderate flowering responses were used. RL was administered from dusk to dawn. Two PGRs, a cytokinin (6-benzyl adenine, BA) and an anti‑ethylene (silver thiosulphate, STS) were applied. Pollen viability was assessed basing on pollen grain diameter, in vitro stainability, in vivo germinability, ovule fertilizability and ploidy level. Treating flowers with RL led to increased pollen diameter from 143.6 to 148.5µm, stainability from 82.9 to 93.0%, ovule fertilizability from 52.0 to 56.9% in treated female flowers and 47.7 to 64.3% on using pollen from treated male flowers. Fruit set increased from 51.5 to 71.8% in RL-treated female flowers and from 53.7 to 71.1% when pollen from RL-treated male flowers was used. Seed set followed a trend similar to that of fruit set. Ploidy level of pollen from RL-treated flowers increased slightly and positively correlated with pollen diameter (R2 = 0.09*), ovule fertilization (R2 = 0.20*), fruit (R2 = 0.59*) and seed set (R2 = 0.60*); but negatively correlated with pollen stainability. Treating flowers with PGR did not affect pollen diameter and stainability, but increased ovule fertilizability from 42.9 to 64.9% and fruit set from 23.2 to 51.9% in treated female flowers and 31.3 to 54.7% when pollen from treated male flowers was used. Application of combined BA+STS on male flowers caused the highest ovule fertilizability, fruit and seed set efficiency, though this varied among study genotypes. These results show that RL and PGR treatment of flowers increases pollen viability and ovule fertilizability. This is important for planning pollination strategies in cassava breeding programmes.

Cassava breeding faces obstacles due to late flowering and poor flower and seed set. The acceleration of breeding processes and the reduction in each cycle’s duration hinge upon efficiently conducting crosses to yield ample progeny for subsequent cycles. Our primary objective was to identify methods that provide tools for cassava breeding programs, enabling them to consistently and rapidly generate offspring from a wide array of genotypes. In greenhouse trials, we examined the effects of the anti-ethylene silver thiosulfate (STS) and the cytokinin benzyladenine (BA). STS, administered via petiole infusion, and BA, applied as an apical spray, combined with the pruning of young branches, significantly augmented the number of flowers. Controls produced no flowers, whereas treatments with pruning plus either BA or STS alone produced an average maximum of 86 flowers per plant, and the combination of pruning, BA and STS yielded 168 flowers per plant. While STS had its primary effect on flower numbers, BA increased the fraction of female flowers from less than 20% to ≥87%, thus increasing the number of progeny from desired parents. Through field studies, we devised an optimal protocol that maintained acceptable levels of phytodamage ratings while substantially increasing seed production per plant compared to untreated plants. This protocol involves adjusting the dosage and timing of treatments to accommodate genotypic variations. As a result, cassava breeding programs can effectively leverage a diverse range of germplasm to develop cultivars with the desired traits.

Background Cassava ( Manihot esculenta Crantz) is staple food and major source of calories for over 500 million people in sub-Saharan Africa. The crop is also a source of income for smallholder farmers, and has increasing potential for industrial utilization. However, breeding efforts to match the increasing demand of cassava are impeded by its inability to flower, delayed or unsynchronized flowering, low proportion of female flowers and high fruit abortions. To overcome these sexual reproductive bottlenecks, this study investigated the effectiveness of using red lights to extend the photoperiod (RLE), as a gateway to enhancing flowering and fruit set under field conditions. Materials and methods Panels of cassava genotypes, with non- or late and early flowering response, 10 in each case, were subjected to RLE from dusk to dawn. RLE was further evaluated at low (LL), medium (ML) and high (HL) red light intensities, at ~ ≤ 0.5; 1.0 and 1.5PFD (Photon Flux Density) in µmol m −2 s −1 respectively. Additionally, the effect of a cytokinin and anti-ethylene as plant growth regulators (PGR) and pruning under RLE treatment were examined. Results RLE stimulated earlier flower initiation in all genotypes, by up to 2 months in the late-flowering genotypes. Height and number of nodes at first branching, particularly in the late-flowering genotypes were also reduced, by over 50%. Number and proportion of pistillate flowers more than doubled, while number of fruits and seeds also increased. Number of branching levels during the crop season also increased by about three. Earlier flowering in many genotypes was most elicited at LL to ML intensities. Additive effects on flower numbers were detected between RLE, PGR and pruning applications. PGR and pruning treatments further increased number and proportion of pistillate flowers and fruits. Plants subjected to PGR and pruning, developed bisexual flowers and exhibited feminization of staminate flowers. Pruning at first branching resulted in higher pistillate flower induction than at second branching. Conclusions These results indicate that RLE improves flowering in cassava, and its effectiveness is enhanced when PGR and pruning are applied. Thus, deployment of these technologies in breeding programs could significantly enhance cassava hybridizations and thus cassava breeding efficiency and impact.

Cassava is a tropical crop that provides daily carbohydrates to more than 800 million people. New cassava cultivars with improved yield, disease resistance, and food quality are critical to end hunger and reduce poverty in the tropics. However, the progress of new cultivar development has been dragged down by difficulties obtaining flowers from desired parental plants to enable designed crosses. Inducing early flowering and increasing seed production are crucial to improving the efficiency of developing farmer-preferred cultivars. In the present study, we used breeding progenitors to evaluate the effectiveness of flower-inducing technology, including photoperiod extension, pruning, and plant growth regulators. Photoperiod extension significantly reduced the time to flowering in all 150 breeding progenitors, especially late-flowering progenitors which were reduced from 6-7 months to 3-4 months. Seed production was increased by using the combination of pruning and plant growth regulators. Combining photoperiod extension with pruning and the PGR 6-benzyladenine (synthetic cytokinin) produced significantly more fruits and seeds than only photoperiod extension and pruning. Another growth regulator, silver thiosulfate, commonly used to block the action of ethylene, did not show a significant effect on fruit or seed production when combined with pruning. The present study validated a protocol for flower induction in cassava breeding programs and discussed factors to consider in implementing the technology. By inducing early flowering and increasing seed production, the protocol helped move one step further for speed breeding in cassava.

Cassava is a staple food crop in the tropics, and is of particular importance in Africa. Recent development of genomic selection technology have improved the speed of cassava breeding; however, cassava flower initiation and development remains a bottleneck. The objectives of the current studies were to elucidate the effect of photoperiod, temperature and their interactions on the time of flowering and flower development in controlled environments, and to use RNA-sequencing to identify transcriptome expression underlying these environmental responses. Compared to a normal tropical day-length of 12 h, increasing the photoperiod by 4 h or decreasing the air temperature from 34/31 to 22°/19°C (day/night) substantially hastened the time to flowering. For both photoperiod and temperature, the environment most favorable for flowering was opposite the one for storage root harvest index. There was a pronounced treatment interaction: at warm day-time temperatures, percent flowering was low, and photoperiod had little effect. In contrast, at cooler temperatures, percent flowering increased, and long-day (LD) photoperiod had a strong effect in hastening flowering. In response to temperature, many differentially expressed genes in the sugar, phase-change, and flowering-time-integrator pathways had expression/flowering patterns in the same direction as in Arabidopsis (positive or negative) even though the effect of temperature on flowering operates in the reverse direction in cassava compared to Arabidopsis. Three trehalose-6-phosphate-synthase-1 (TPS1) genes and four members of the SPL gene family had significantly increased expression at cool temperature, suggesting sugar signaling roles in flower induction. In response to LD photoperiod, regulatory genes were expressed as in Arabidopsis and other LD flowering plants. Several hormone-related genes were expressed in response to both photoperiod and temperature. In summary, these findings provide insight on photoperiod and temperature responses and underlying gene expression that may assist breeding programs to manipulate flowering for more rapid crop improvement.

Cassava, a tropical storage-root crop, is a major source of food security for millions in the tropics. Cassava breeding, however, is hindered by the poor development of flowers and a low ratio of female flowers to male flowers. To advance the understanding of the mechanistic factors regulating cassava flowering, combinations of plant growth regulators (PGRs) and pruning treatments were examined for their effectiveness in improving flower production and fruit set in field conditions. Pruning the fork-type branches, which arise at the shoot apex immediately below newly formed inflorescences, stimulated inflorescence and floral development. The anti-ethylene PGR silver thiosulfate (STS) also increased flower abundance. Both pruning and STS increased flower numbers while having minimal influence on sex ratios. In contrast, the cytokinin benzyladenine (BA) feminized flowers without increasing flower abundance. Combining pruning and STS treatments led to an additive increase in flower abundance; with the addition of BA, over 80% of flowers were females. This three-way treatment combination of pruning+STS+BA also led to an increase in fruit number. Transcriptomic analysis of gene expression in tissues of the apical region and developing inflorescence revealed that the enhancement of flower development by STS+BA was accompanied by downregulation of several genes associated with repression of flowering, including homologs of TEMPRANILLO1 (TEM1), GA receptor GID1b, and ABA signaling genes ABI1 and PP2CA. We conclude that flower-enhancing treatments with pruning, STS, and BA create widespread changes in the network of hormone signaling and regulatory factors beyond ethylene and cytokinin.

Flowering in cassava is closely linked with branching. Early-flowering genotypes branch low and abundantly. Although farmers prefer late flowering genotypes because of their erect plant architecture, their usefulness as progenitors in breeding is limited by their low seed production. In general, the first inflorescence aborts in cassava. Preventing this abortion would result in early production of seeds and make cassava breeding more efficient. The objective of this study was to assess if pruning young branches prevents the abortion of first inflorescences and promotes early fruit and seed set. Four genotypes with early, late, very late, and no flowering habits were grown under an extended photoperiod (EP) or normal dark night conditions (DN). Additional treatments included pruning young branches at the first or second flowering event and spraying (or not) benzyladenine (BA) after pruning. One genotype failed to flower and was not considered further. For the remaining genotypes, EP proved crucial to induce an earlier flowering, which is a pre-requisite for pruning. Total production of seeds in EP plots was 2,971versus150 in DN plots. For plants grown under EP, the average number of seeds per plant without pruning was 3.88, whereas those pruned produced 17.60 seeds per plant. Pruning at the first branching event led to higher number of seeds per plant (26.25) than pruning at the second flowering event (8.95). In general, applying BA was beneficial (38.52 and 13.98 seeds/plant with or without spraying it, respectively). The best combination of treatments was different for each genotype. Pruning young branches and applying BA in the first flowering event not only prevented the abortion of inflorescences but also induced the feminization of male flowers into hermaphrodite or female-only flowers. The procedures suggested from this study (combining EP, pruning young branches, and spraying BA), allowed the production of a high number of seeds from erect cassava genotypes in a short period. The implementation of these procedures will improve the breeding efficiency in cassava.

Cassava, a tropical storage-root crop, is a major source of food security for millions in the tropics. Cassava breeding however is hindered by the poor development of flowers and female flowers in particular, since flower development is strongly skewed towards male flowers. Our objectives were to test plant growth regulator and pruning treatments for their effectiveness in field conditions in improving flower production and fruit set in cassava. Pruning the fork type branches that arise at the shoot apex immediately below newly formed inflorescences stimulated inflorescence and floral development. The anti-ethylene silver thiosulfate (STS) also increased flower abundance. Both pruning and STS increased flower numbers without influencing sex ratios. In contrast, the cytokinin benzyladenine (BA) feminized flowers without increasing flower abundance. Combining pruning and STS treatments led to an additive increase in flower abundance; with the addition of BA, over 80% of flowers were females. This three-way treatment combination of pruning+STS+BA also led to an increase in fruit development. Transcriptomic analysis of gene expression in tissues of the apical region and developing inflorescence revealed that the enhancement of female flower development by STS+BA was accompanied by the downregulation in of several genes associated with repression of flowering, including Tempranillo 1 (TEM1), GA receptor GID1b, and ABA signaling genes ABI1 and PP2CA. We conclude that treatments with pruning, STS and BA create widespread changes on the network of hormone signaling and regulatory factors beyond ethylene and cytokinin. ### Competing Interest Statement The authors have declared no competing interest.

Erect plant architecture is preferred by farmers but results in late and scarce flowering, which slows down breeding considerably. Inducing earlier and abundant flowering in crossing nurseries (involving erect genotypes) is a key objective for cassava and was the subject of this study. Five genotypes with contrasting flowering behavior were grown under dark night (DN) and extended photoperiod (EP) conditions for three seasons. EP was achieved with different red light emitting diodes (LEDs) with 625–635 nm wavelength all night long or through night-breaks. EP reduced height and number of days to first branching, particularly in non- or late-flowering genotypes. A minimum of 0.02 μmol m−2 s−1 was required to elicit earlier flowering in plants illuminated all night. Early results using five genotypes were validated across 116 genotypes planted in a crossing nursery. EP promoted earlier flowering in erect-plant genotypes but reduced the number of branching events in early flowering genotypes to some extent. 50W LED lamps, fixed at 3 m above ground in a 4.5 m grid, proved to be a practical approach to extend photoperiod in breeding nurseries. Night breaks also proved effective, thus opening the possibility of using solar panels where electricity is not available.

Cassava is a starch-storing root crop that is an important source of dietary energy in tropical regions of the world. Genetic improvement of cassava by breeding is hindered by late flowering and sparse flower production in lines that are needed as parents. To advance understanding of regulatory mechanisms in cassava, this work sought to identify and characterize homologs of the FLOWERING LOCUS T (FT) gene. Ten members of the phosphatidylethanolamine-binding protein gene family, to which FT belongs, were obtained from the cassava genome database. Phylogenetic and sequence analysis of these proteins was used to identify two putative FT homologs which had amino acid sequences at key positions in accordance with those predicted for functional FTs. Expression of these ten genes was determined in mature leaves, immature leaves, flower buds, fibrous roots, storage roots and stem. The FT transcripts were expressed in mature leaves, as expected for their possible role in leaf-to-apical meristem signaling. In growth chamber studies, plants flowered earlier in long-day photoperiod than in short-day photoperiod. Expression studies indicated that while MeFT1 was expressed in leaves without a clear-cut photoperiod response, MeFT2 was expressed in a photoperiod-dependent manner, consistent with its involvement in photoperiodic control of flowering. In growth chambers that subjected plants to a range of temperatures from 22 to 34 °C, flowering was delayed by warmer temperatures although MeFT1 and MeFT2 expression declined in only one genotype, indicating other factors regulate this response. The earliest flowering genotype, IBA980002, had high levels of MeFT1 and MeFT2 expression, suggesting that both homologs contribute to earliness of this genotype.

Cassava, which produces edible starchy roots, is an important staple food for hundreds of millions of people in the tropics. Breeding of cassava is hampered by its poor flower production, flower abortion, and lack of reproductive prolificacy. The current work determined that ethylene signalling affects floral development in cassava and that the anti-ethylene plant growth regulator silver thiosulfate (STS) mitigates the effects of ethylene on flower development. STS did not affect the timing of flower initiation, but improved early inflorescence and flower development as well as flower longevity such that flower numbers were increased. STS did not affect shoot and storage root growth. Studies of silver accumulation and treatment localization support the hypothesis that the beneficial effects of STS are confined to tissues of the shoot apex. The most effective timing of application was before inflorescence appearance extending to post-flower appearance. Based on this work a recommended protocol for STS use was developed. This work has the potential to improve methods for enhancing cassava flower development in breeding nurseries and thereby synchronize flowering of desired parents and enable the production of abundant progeny of desired crosses.

ABSTRACT Onion ( Allium cepa L.) lines resistant to Botrytis leaf blight (BLB) were produced by backcrossing the Bs1 BLB resistance of Allium roylei Stearn into cultivated onion. This study evaluated important issues for the commercial use of BLB‐resistant onions: the relative degree of BLB control possible when Bs1 is homozygous rather than heterozygous, the control achieved against multiple pathogen isolates, and the potential for negative traits associated with transfer of Bs1 . Homozygous BLB‐resistant onions tested in inoculated growth chamber screens with five distinct Botrytis squamosa Walker isolates and in naturally infested regional field trials showed significantly lower BLB symptoms than susceptible commercial hybrid controls. F 1 hybrids heterozygous for the Bs1 gene showed levels of BLB symptoms between those of the homozygous resistant and susceptible controls, confirming previous reports of partial dominance. The difference in BLB control in plants heterozygous for Bs1 compared with homozygous plants was more noticeable under heavier disease pressure. Hybrids heterozygous for Bs1 had similar level of BLB control in most of the field trials as their parental homozygous BLB‐resistant lines, indicating that either heterozygous or homozygous BLB resistant hybrids could be used commercially for BLB control. There were no differences in the degree of pathogenicity across the five B. squamosa isolates, which were genetically different based on polymorphism for DNA markers, or among the naturally occurring pathogens present at the six different field locations used. No obvious association was noted between presence of the Bs1 resistance gene with unfavorable characteristics such as reduced bulb size or yield.

Production of double haploid (DH) lines is a rapid method of obtaining completely homozygous inbred lines for numerous crops; however, practical trials testing the use of DH onion lines have been limited. DH onion lines were produced from diverse highly heterozygous material in development within the Cornell onion breeding program. These DH lines were evaluated in multiple replicated trials in onion fields in New York as lines and as parents of hybrids to assess the impact and commercial potential of DH onions. Twenty DH onion lines were compared with open-pollinated cultivars developed from the same source germplasm and with a commercial hybrid over two years. The vegetative vigor of the DH lines was comparable to that of related open-pollinated cultivars, showing minimal, if any, inbreeding depression. Two sets of hybrids were produced using the DH lines as males and two different females that are the female parents of the commercial hybrid controls. Therefore, hybrids in each set are half-sibs, and any performance differences are attributable to their DH male lines. In four replicated trials of these hybrids and controls, the experimental hybrids were either not significantly different or significantly better in measures of vegetative vigor compared with conventional half-sib hybrid controls. The vegetative vigor of DH lines, and their derived hybrids, might result from selection of plantlets without deleterious sublethal genes during gynogenesis. The shortened development time and equivalent quality of DH lines compared with the traditionally bred onion inbreds warrant their use. The increased vigor in hybrid combination could be an additional benefit for onion breeding strategies.