Mining the A.E. Watkins Wheat Landrace Collection for Variation in Starch Digestibility Using a New High-Throughput Assay.

Breeding for less digestible starch in wheat can improve the health impact of bread and other wheat foods. The application of forward genetic approaches has lately opened opportunities for the discovery of new genes that influence the digestibility of starch, without the burden of detrimental effects on yield or on pasta and bread-making quality. In this study we developed a high-throughput in vitro starch digestibility assay (HTA) for use in forward genetic approaches to screen wheat germplasm. The HTA was validated using standard maize and wheat starches. Using the HTA we measured starch digestibility in hydrothermally processed flour samples and found wide variation among 118 wheat landraces from the A. E. Watkins collection and among eight elite UK varieties (23.5 to 39.9% and 31.2 to 43.5% starch digested after 90 min, respectively). We further investigated starch digestibility in fractions of sieved wholemeal flour and purified starch in a subset of the Watkins lines and elite varieties and found that the matrix properties of flour rather than the intrinsic properties of starch granules conferred lower starch digestibility.

Loss of starch synthase IIIa changes starch molecular structure and granule morphology in grains of hexaploid bread wheat.

Starch synthase III plays a key role in starch biosynthesis and is highly expressed in developing wheat grains. To understand the contribution of SSIII to starch and grain properties, we developed wheat ssIIIa mutants in the elite cultivar Cadenza using in silico TILLING in a mutagenized population. SSIIIa protein was undetectable by immunoblot analysis in triple ssIIIa mutants carrying mutations in each homoeologous copy of ssIIIa (A, B and D). Loss of SSIIIa in triple mutants led to significant changes in starch phenotype including smaller A-type granules and altered granule morphology. Starch chain-length distributions of double and triple mutants indicated greater levels of amylose than sibling controls (33.8% of starch in triple mutants, and 29.3% in double mutants vs. 25.5% in sibling controls) and fewer long amylopectin chains. Wholemeal flour of triple mutants had more resistant starch (6.0% vs. 2.9% in sibling controls) and greater levels of non-starch polysaccharides; the grains appeared shrunken and weighed ~ 11% less than the sibling control which was partially explained by loss in starch content. Interestingly, our study revealed gene dosage effects which could be useful for fine-tuning starch properties in wheat breeding applications while minimizing impact on grain weight and quality.

Effect of high-amylose starch branching enzyme II wheat mutants on starch digestibility in bread, product quality, postprandial satiety and glycaemic response.

High-amylose starch branching enzyme II (sbeII) mutant wheat has potential to be low-glycaemic compared to conventional wheat; however, the effects of bread made from sbeII wheat flour on glycaemic response and product quality require investigation. We report the impact of white bread made from sbeII wheat flour on in vitro starch digestibility and product quality, and on postprandial glycaemia in vivo, compared to an isoglucidic wild-type (WT) control white bread. Starch in sbeII bread was ∼20% less susceptible to in vitro amylolysis leading to ∼15% lower glycaemic response measured in vivo, compared to the WT control bread, without major effects on bread appearance or texture, measured instrumentally. Despite the early termination of the in vivo intervention study due to the COVID-19 outbreak (n = 8 out of 19), results from this study indicate that sbeII wheat produces bread with lower starch digestibility than conventional white bread.

Effect of semolina pudding prepared from starch branching enzyme IIa and b mutant wheat on glycaemic response in vitro and in vivo: a randomised controlled pilot study.

Refined starchy foods are usually rapidly digested, leading to poor glycaemic control, but not all starchy foods are the same. Complex carbohydrates like resistant starch (RS) have been shown to reduce the metabolic risk factors for chronic diseases such as hyperglycaemia and overweight. The aim of the project was to develop a semolina-based food made from a starch branching enzyme II (sbeIIa/b-AB) durum wheat mutant with a high RS content and to measure its glycaemic index using a double-blind randomised pilot study. We report here the amylose, RS and non-starch polysaccharide concentration of raw sbeIIa/b-AB and wild-type control (WT) semolina. We measured RS after cooking to identify a model food for in vivo testing. Retrograded sbeIIa/b-AB semolina showed a higher RS concentration than the WT control (RS = 4.87 ± 0.6 g per 100 g, 0.77 ± 0.34 g per 100 g starch DWB, respectively), so pudding was selected as the test food. Ten healthy participants consumed ∼50 g of total starch from WT and sbeIIa/b-AB pudding and a standard glucose drink. Capillary blood glucose concentrations were measured in the fasting and postprandial state (2 h): incremental area-under-the-curve (iAUC) and GI were calculated. We found no evidence of difference in GI between sbeIIa/b-AB pudding and the WT control, but the starch digestibility was significantly lower in sbeIIa/b-AB pudding compared to the WT control in vitro (C90 = 33.29% and 47.38%, respectively). Based on these results, novel sbeIIa/b-AB wheat foods will be used in future in vivo studies to test the effect of different RS concentrations and different food matrices on glycaemia.