Mark Wohlers

Genomic Analysis of the Kiwifruit Pathogen Pseudomonas syringae pv. actinidiae Provides Insight into the Origins of an Emergent Plant Disease

The origins of crop diseases are linked to domestication of plants. Most crops were domesticated centuries – even millennia – ago, thus limiting opportunity to understand the concomitant emergence of disease. Kiwifruit (Actinidia spp.) is an exception: domestication began in the 1930s with outbreaks of canker disease caused by P. syringae pv. actinidiae (Psa) first recorded in the 1980s. Based on SNP analyses of two circularized and 34 draft genomes, we show that Psa is comprised of distinct clades exhibiting negligible within-clade diversity, consistent with disease arising by independent samplings from a source population. Three clades correspond to their geographical source of isolation; a fourth, encompassing the Psa-V lineage responsible for the 2008 outbreak, is now globally distributed. Psa has an overall clonal population structure, however, genomes carry a marked signature of within-pathovar recombination. SNP analysis of Psa-V reveals hundreds of polymorphisms; however, most reside within PPHGI-1-like conjugative elements whose evolution is unlinked to the core genome. Removal of SNPs due to recombination yields an uninformative (star-like) phylogeny consistent with diversification of Psa-V from a single clone within the last ten years. Growth assays provide evidence of cultivar specificity, with rapid systemic movement of Psa-V in Actinidia chinensis. Genomic comparisons show a dynamic genome with evidence of positive selection on type III effectors and other candidate virulence genes. Each clade has highly varied complements of accessory genes encoding effectors and toxins with evidence of gain and loss via multiple genetic routes. Genes with orthologs in vascular pathogens were found exclusively within Psa-V. Our analyses capture a pathogen in the early stages of emergence from a predicted source population associated with wild Actinidia species. In addition to candidate genes as targets for resistance breeding programs, our findings highlight the importance of the source population as a reservoir of new disease.

Identifying the Chemical Composition Related to the Distinct Aroma Characteristics of New Zealand Sauvignon blanc Wines

A comprehensive set of aroma compounds was quantified in 79 Sauvignon blanc wines from different international producers. Emphasis was given to understanding the chemical differences that can explain the unique character of Marlborough Sauvignon blanc. Quantification revealed the potential importance of several volatile compounds in addition to the polyfunctional mercaptans and methoxypyrazines already known to be important to the aroma of Sauvignon blanc wines. Multivariate statistical approaches, including canonical variate analysis, classification tree and partial least square analysis, established correlations between the chemical and the sensory profiles of the wines. A significant role of 3-mercaptohexyl acetate and 3-mercaptohexanol in the unique tropical, fruity character of Marlborough Sauvignon blanc wines was demonstrated, together with important variations in their concentrations, pointing to different styles even within the Marlborough wines.

Stability of antioxidants in an apple polyphenol-milk model system

The stability of antioxidants in an apple polyphenol-milk model system was examined. The model system consisted of skim milk fortified with pH-neutralised apple polyphenols (AP, 0-200mg per 100ml milk), with or without ascorbic acid (100mg per 100ml milk). Physical and chemical changes were evaluated after thermal treatment (120°C, 5min) and oxidative storage (20°C and 38°C, up to 12 weeks). Antioxidant capacity was determined using both oxygen radical absorbance capacity (ORAC) assay and ferric reducing antioxidant power (FRAP) assay. Significant antioxidant capacity was detected in the presence of milk. Antioxidant capacity was retained during thermal treatment but decreased slowly during storage. The concentration of ascorbic acid decreased rapidly, and was close to zero after 2-week storage at 38°C or 10-week storage at 20°C. The brownness of the polyphenol-milk system increased over storage duration of 0-12 weeks; this effect was retarded by the addition of ascorbic acid. This high polyphenol-milk has demonstrated good physical stability.

Evaluation of key odorants in sauvignon blanc wines using three different methodologies.

In this study three different approaches were employed to identify key odorants in Sauvignon blanc wines. First, the concentrations of the odorants were compared to their respective aroma detection thresholds. The resulting odor activity values (OAV) were transformed into a normalized and weighted measure that allows the aroma profiles of different wines to be compared and the contribution of a single aroma in a complex mixture to be evaluated. Based on their OAV, 3-mercaptohexanol and 3-mercaptohexyl acetate were the two most important aroma compounds in many Marlborough Sauvignon blanc wines. Due to limitations with the OAV approach, the study was extended to include aroma extract dilution analysis (AEDA), which revealed that β-damascenone, together with the varietal thiols, esters, and higher alcohols, are key odorants in Sauvignon blanc wines. The final approach undertaken was aroma reconstitution and omission tests using a deodorized wine base and the creation of a model Marlborough Sauvignon blanc. Single compounds and groups of compounds were omitted from the model to study their impact on the sensory properties of the model wine. Reconstitution and omission confirmed that varietal thiols, esters, terpenes, and β-damascenone are all important contributors to Sauvignon blanc aroma. The methoxypyrazines showed an important but relatively low impact in all three of the approaches undertaken in this study.

Metabolic analysis of kiwifruit (Actinidia deliciosa) berries from extreme genotypes reveals hallmarks for fruit starch metabolism

Tomato, melon, grape, peach, and strawberry primarily accumulate soluble sugars during fruit development. In contrast, kiwifruit (Actinidia Lindl. spp.) and banana store a large amount of starch that is released as soluble sugars only after the fruit has reached maturity. By integrating metabolites measured by gas chromatography–mass spectrometry, enzyme activities measured by a robot-based platform, and transcript data sets during fruit development of Actinidia deliciosa genotypes contrasting in starch concentration and size, this study identified the metabolic changes occurring during kiwifruit development, including the metabolic hallmarks of starch accumulation and turnover. At cell division, a rise in glucose (Glc) concentration was associated with neutral invertase (NI) activity, and the decline of both Glc and NI activity defined the transition to the cell expansion and starch accumulation phase. The high transcript levels of β-amylase 9 (BAM9) during cell division, prior to net starch accumulation, and the correlation between sucrose phosphate synthase (SPS) activity and sucrose suggest the occurrence of sucrose cycling and starch turnover. ADP-Glc pyrophosphorylase (AGPase) is identified as a key enzyme for starch accumulation in kiwifruit berries, as high-starch genotypes had 2- to 5-fold higher AGPase activity, which was maintained over a longer period of time and was also associated with enhanced and extended transcription of the AGPase large subunit 4 (APL4). The data also revealed that SPS and galactinol might affect kiwifruit starch accumulation, and suggest that phloem unloading into kiwifruit is symplastic. These results are relevant to the genetic improvement of quality traits such as sweetness and sugar/acid balance in a range of fruit species.

Characterization and Quantification of Anthocyanins in Red Kiwifruit (Actinidia spp.)

Red-fleshed fruit occur in a small number of distantly related taxa in different sections of the genus Actinidia (kiwifruit). We describe and identify the anthocyanin profile of fruit of several Actinidia species. Differences in the relative amounts of cyanidin- and delphinidin-based anthocyanins determine whether the fruit appear red or purple. Cyanidin derivatives have been found in all Actinidia species that contain anthocyanins, whereas delphinidin derivatives are limited to two taxa: A. melanandra and A. arguta var. purpurea . The fruit of these not only contain a wider range of anthocyanins, but they also have greater concentrations. Anthocyanins of most Actinidia species are usually conjugated with either xylosyl-galactose or galactose, whereas A. deliciosa anthocyanins are conjugated with glucose and galactose.

Environmental control of branching in petunia.

Nutrient availability and light quality regulate branching by the production and perception of strigolactone. Plants alter their development in response to changes in their environment. This responsiveness has proven to be a successful evolutionary trait. Here, we tested the hypothesis that two key environmental factors, light and nutrition, are integrated within the axillary bud to promote or suppress the growth of the bud into a branch. Using petunia (Petunia hybrida) as a model for vegetative branching, we manipulated both light quality (as crowding and the red-to-far-red light ratio) and phosphate availability, such that the axillary bud at node 7 varied from deeply dormant to rapidly growing. In conjunction with the phenotypic characterization, we also monitored the state of the strigolactone (SL) pathway by quantifying SL-related gene transcripts. Mutants in the SL pathway inhibit but do not abolish the branching response to these environmental signals, and neither signal is dominant over the other, suggesting that the regulation of branching in response to the environment is complex. We have isolated three new putatively SL-related TCP (for Teosinte branched1, Cycloidia, and Proliferating cell factor) genes from petunia, and have identified that these TCP-type transcription factors may have roles in the SL signaling pathway both before and after the reception of the SL signal at the bud. We show that the abundance of the receptor transcript is regulated by light quality, such that axillary buds growing in added far-red light have greatly increased receptor transcript abundance. This suggests a mechanism whereby the impact of any SL signal reaching an axillary bud is modulated by the responsiveness of these cells to the signal.

No difference in fecal levels of bacteria or short chain fatty acids in humans, when consuming fruit juice beverages containing fruit fiber, fruit polyphenols, and their combination ☆

This study examined the effect of a Boysenberry beverage (750 mg polyphenols), an apple fiber beverage (7.5 g dietary fiber), and a Boysenberry plus apple fiber beverage (750 mg polyphenols plus 7.5 g dietary fiber) on gut health. Twenty-five individuals completed the study. The study was a placebo-controlled crossover study, where every individual consumed 1 of the 4 treatments in turn. Each treatment phase was 4-week long and was followed by a 2-week washout period. The trial beverages were 350 g taken in 2 doses every day (ie, 175 mL taken twice daily). The hypothesis for the study was that the combination of polyphenols and fiber would have a greater benefit on gut health than the placebo product or the fiber or polyphenols on their own. There were no differences in fecal levels of total bacteria, Bacteroides-Prevotella-Porphyromonas group, Bifidobacteriumspecies, Clostridium perfringens, or Lactobacillus species among any of the treatment groups. Fecal short chain fatty acid concentrations did not vary among treatment groups, although prostaglandin E2 concentrations were higher after consumption of the Boysenberry juice beverage. No significant differences were found in quantitative measures of gut health between the Boysenberry juice beverage, the apple fiber beverage, the Boysenberry juice plus apple fiber beverage, and the placebo beverage.

Red-foliaged apples affect the establishment, growth, and development of the light brown apple moth, Epiphyas postvittana

Anthocyanins have diverse roles in plant reproduction and in response to both abiotic and biotic stress. By over‐expressing the apple MYB transcription factor, MYB10, we have generated apple trees, Malus × domestica Borkh. (Rosaceae), with highly pigmented red foliage due to the presence of high concentrations of cyanidin‐based anthocyanins. In this study, we investigated the impact of the high anthocyanic apple leaves on the behaviour, feeding, and life cycle parameters of the light brown apple moth, Epiphyas postvittana (Walker) (Lepidoptera: Tortricidae), a widespread herbivorous pest of apples. Behaviour of 1‐ to 3‐day‐old larvae towards red anthocyanin‐rich and green apple leaves was studied in choice and no‐choice trials, and survival, growth, and development of larvae feeding on red and green apple leaves from the neonate to pupation were compared. First‐instar E. postvittana showed a statistically significant preference for green over red leaves from three transgenic red apple lines and a preference for the less red old leaves over the bright red new leaves. This was significantly different from their behaviour towards wild type green leaves, where new leaves were selected over old. In complete darkness, first‐instar E. postvittana showed no significant preference for the green leaves, indicating that visual cues were important. Although larval survival was not greatly affected by feeding on the red foliage, larval growth and development were significantly affected by feeding on leaves from some of the transgenic red apple lines. Liquid chromatography‐mass spectrometry analysis of leaf phenolics showed that both anthocyanins and quercetin glycosides were significantly raised in the red transgenic lines compared with wild types.

Dry matter content and fruit size affect flavour and texture of novel Actinidia deliciosa genotypes.

BACKGROUND Previous studies with commercial kiwifruit cultivars have demonstrated that the taste of fruit with higher dry matter content (DM) is more liked by consumers. A unique replicated trial of kiwifruit genotypes (10 high/low DM × small/large-fruited genotypes) has provided an opportunity to consider how the genetic propensity for a kiwifruit to accumulate DM affects fruit flavour and texture. In the present study, eating-ripe fruit from each of the genotypes were assessed using a trained sensory panel and the relationships between these sensory attributes and fresh weight, DM, flesh firmness and soluble solids content (SSC) were explored. RESULTS The genotypes provided a diversity of flavour and texture attributes, each of which varied in perceived intensity of the sensory experience. High-DM genotypes had higher SSC and were perceived as sweeter than low-DM genotypes. Sweet taste was closely associated with the perception of the tropical flavour and high-DM genotypes were found to have more tropical notes. Fruit size was associated with fruit texture, and small fruit were characterised by a firmer and more fibrous core. Large high-DM fruit were perceived as juicier than those of all other genotypes. CONCLUSIONS Genotypes were perceived differently from one another, and differences in fruit size and DM content were reflected in fruit sensorial properties. This study is unique in demonstrating interactions between fruit size, DM and sensory properties. These findings could be relevant not only to kiwifruit but to fruiting crop breeders in general, because of the demonstrated potential for effects of fruit size and DM content on sweetness, flavour and fruit texture.