Table beet is a minor vegetable crop with strong economic importance in Wisconsin, which led 2017 U.S. table beet production with over 1600 ha harvested. The United States table beet harvest serves both processing (52%) and fresh (48%) markets (U.S. Department of Agriculture, 2017) with both standard red table beets and lesser volumes of novel cultivars (Goldman and Navazio, 2003). Flavor is critical to consumer acceptance of horticultural crops, so while different table beet flavor profiles may be appropriate for different markets (Dawson and Healy, 2018) and preparation methods (Bach et al., 2015), achieving consumer-accepted flavor is essential to table beet marketability. Four chemical compounds are documented to be associated with table beet flavor: geosmin, sucrose, oxalate, and saponins. Earthy aroma, conferred by the volatile terpenoid geosmin (Gerber, 1967), is identified as the signature flavor of table beet (Goldman and Navazio, 2003) but can be unpalatable in excess (Tyler et al., 1979). Sucrose, the major saccharide molecule in table beet root (Bach et al., 2015), contributes sweet flavor and is commonly measured as TDS. Oxalate is present in table beet (Freidig and Goldman, 2011) and has been associated with abrasive sensory properties in other plants (Korth et al., 2006; Salinas et al., 2001). Saponins, which confer bitter flavor in other crops, have also been identified in table beet (Mikołajczyk-Bator et al., 2016). The present research seeks to understand the genetic control and genotype × environment interactions associated with geosmin concentration and TDS in table beet, to facilitate breeding for desired table beet flavor.
The earthy flavor in table beet is conferred by geosmin (trans-1,10-dimethyl-trans-9-decalol), the volatile organic compound that imbues moist earth with its distinctive aroma (Marshall and Hochstetler, 1968). Geosmin is synthesized by a diverse group of bacteria, cyanobacteria, and fungi; Streptomyces bacteria are recognized as prominent producers of geosmin (Spiteller et al., 2002). Geosmin is known to occur in several Amaranthaceae family members, including spinach (Spinacea oleracea), swiss chard (B. vulgaris ssp. cicla), and table beet (Acree and Lee, 1976), while dehydrogeosmin, a compound that can assume a conformation identical to geosmin, has been identified in six genera of the Cactaceae family (Schlumpberger et al., 2004). Humans are exceedingly sensitive to geosmin and able to detect the molecule at concentrations as low as 10 to 20 ng·L−1 water (Tyler et al., 1979). Geosmin is present in drinking water, wine, beer, and various foods, often rendering the affected water or foodstuff unpalatable (Buttery et al., 1976; Darriet et al., 2000; Frisvad et al., 1997). In a sensory study using table beet juice, geosmin conferred characteristic earthy flavor up to a 5.8 μg·L−1 threshold; geosmin concentration above this level was identified by consumers as “too earthy to be beet-like,” while very low geosmin concentration (0.36 μg·L−1) was perceived as lacking characteristic table beet flavor (Tyler et al., 1979). In addition, earthy flavor is anecdotally cited as a reason that consumers avoid table beets (Lu et al., 2003b). Thus, geosmin may be perceived as either pleasing or undesirable, depending on its concentration and the context in which it occurs.
The microbial biosynthesis of geosmin begins with precursor molecule farnesyl diphosphate (FPP). In the presence of a single, bifunctional geosmin synthase enzyme and Mg2+ cofactors, FPP is cyclized into an 85:15 mixture of germacradienol and germacrene D. Next, a cyclization-fragmentation reaction converts the germacradienol fraction into geosmin. The presence of divalent cations Cu2+ and Fe2+ decreases geosmin synthesis in vitro (Jiang et al., 2006) and in Streptomyces halstedii (Schrader and Blevins, 2001), perhaps due to competition with Mg2+ for access to geosmin precursor compounds. A geosmin synthase gene has been found in Streptomyces coelicolor (Cane and Watt, 2003) and several other microbial species (Giglio et al., 2008; Singh et al., 2009).
In 2003, Lu et al. developed a headspace solid-phase microextraction (HSPME) protocol for gas chromatography–mass spectrometry (GC–MS) to quantify geosmin concentration within a table beet slurry matrix. This methodology facilitated inquiry into three possible hypotheses regarding the presence of geosmin in table beet. First, table beet plants might associate preferentially with geosmin-producing microbes, perhaps via genotype-specific root exudates. Geosmin is known to accumulate in and directly under the root epidermis in table beet (Lu et al., 2003a; Tyler et al., 1978), lending initial support to this hypothesis. However, the presence of geosmin in table beet tissue grown in autoclaved soil (Freidig and Goldman, 2014) and sterile tissue culture (Maher and Goldman, 2018) refutes it. Second, geosmin could be produced within table beet tissue by endophytic microbes. However, genomic probes of table beet tissue grown in sterile culture revealed no obvious prokaryotic sequences other than those present in chloroplast DNA (Maher and Goldman, 2018). Thus, evidence accrues for a third possibility: that of endogenous production of geosmin by table beet plants. In support of this possibility—but not expressly in contradiction to the others—table beet populations were responsive to recurrent selection for high and low geosmin concentration, with realized heritabilities of 0.23 and 0.7 for high and low geosmin populations, respectively (Maher and Goldman, 2017).
To search for a geosmin synthase gene in B. vulgaris, Maher (2017) queried the sugar beet (B. vulgaris ssp. vulgaris) reference genome using Position-Specific Iterative Basic Local Alignment Search Tool (PSI-BLAST) for proteins with similar three-dimensional structure as the S. coelicolor geosmin synthase gene, yielding one predicted linalool-synthase-like protein and two hypothetical proteins. The latter two proteins—a terpene synthase and an isoprenoid synthase—are located adjacent to one another on chromosome 8, and both appear to have two functional domains, as does geosmin synthase (Maher, 2017). While these proteins have not been proven to synthesize geosmin from precursor molecules, their existence suggests a mechanism by which geosmin could be produced endogenously via the B. vulgaris genome.
Sweet flavor is another key sensory attribute of table beet, and sugars occur in table beet mainly as sucrose, although small amounts of fructose and glucose are also present (Bach et al., 2015). Sucrose concentration in sugar beet, which shares both species and subspecies with table beet, is highly heritable, quantitatively controlled, and directly related to total root dry mass. Sucrose content in sugar beet has been increased successfully by selecting for root specific gravity—proportionally higher dry matter and lower water content—and for a higher root:shoot biomass ratio (McGrath and Panella, 2018). Narrow-sense heritability (h2) for percent sugar content was estimated at 0.60 in a testcross population of 924 sugar beet genotypes (Würschum et al., 2011). Five quantitative trait loci (QTL) have been associated with sucrose content in sugar beet (Schneider et al., 2002), and several seemed to colocalize with QTL for water content in a preliminary study (Trebbi and McGrath, 2007). Bidirectional selection for TDS in high pigment table beet populations, using an index integrating TDS and betalain pigment concentration, showed moderate response to upward selection and limited response to downward selection, perhaps because of the essential role of sucrose metabolites in the betalain synthesis pathway (Goldman et al., 1996; Wolyn and Gabelman, 1990). Thus, while sucrose concentration in B. vulgaris is under substantial genetic control, environmental factors also contribute to this phenotype.
Lay garden literature suggests that table beet flavor is superior in soils with high magnesium content (High Mowing Organic Seeds, 2016). While soil Mg2+ deficiency is known to inhibit root sugar accumulation in sugar beet (Hermans et al., 2004), perhaps due to the central role of magnesium ions in sucrose-producing chlorophyll molecules (Shaul, 2002), it is plausible that Mg2+ deficiency also limits geosmin synthesis. That is, Mg2+ is an essential cofactor in microbial geosmin biosynthesis, and if a hypothetical B. vulgaris geosmin synthase enzyme also required Mg2+, soil magnesium concentration also could affect geosmin concentration in table beet. In soil, Mg2+ competes with other cations for space on the negatively charged cation exchange complex; after entering plant roots through both apoplasmic and symplasmic pathways, it is loaded into xylem cells and unloaded at ubiquitous locations in plant shoots. Within each plant shoot cell, a quantity of Mg2+ is complexed with adenosine triphosphate (ATP), but the vacuole functions to sequester free Mg2+, tightly regulating its concentration within the cytosol (Shaul, 2002). In sugar beet plants, supplemental magnesium fertilization has been shown to significantly increase in planta Mg2+ concentration (Durrant and Draycott, 1971; Zengin et al., 2009), root yield (Grzebisz, 2013; Zengin et al., 2009), and nitrogen uptake efficiency (Grzebisz, 2013). Effects on yield were most significant when magnesium fertilizer was added to Mg2+-deficient soils with low cation exchange capacity (CEC) (Grzebisz, 2013; Zengin et al., 2009). Magnesium fertilization of sugar beet caused decreased in planta calcium concentration (Durrant and Draycott, 1971), showing that Mg2+ and Ca2+ act as competing cations within sugar beet tissue. The effect of magnesium fertilization on geosmin concentration in Beta species has not been studied.
Flavor compounds in horticultural crops are known to vary with genotype, environment, and their interaction, but this variation seems to be specific to crop, compound, and environment (Cebolla-Cornejo et al., 2011; Crespo et al., 2010; Duckham et al., 2001, 2002; Jensen et al., 1999). Previous genotype × environment studies of TDS in table beet showed strong environmental effects and limited realized heritability (Goldman et al., 1996), but these estimates were attained using genotypes selected for high pigment concentration rather than commercial horticultural use. In a study of geosmin concentration in table beet, Freidig and Goldman (2014) found highly significant genotype and non-crossover genotype × environment interaction effects, plus a moderately significant environmental effect. The genotype × environment interaction reflected significantly different geosmin concentration between field and greenhouse environments within cultivar, perhaps due to the smaller size of the greenhouse-grown roots. Indeed, an early study analyzing whole-root samples of ‘Ruby Queen’ table beet found significantly higher geosmin concentration in smaller roots than in larger roots (Tyler et al., 1978), likely due to incorporation of proportionally more epidermal tissue to core root tissue in homogenate made from small roots. Freidig and Goldman (2014) used a sampling method designed to standardize the proportion of epidermal to core root tissue in each sample, but because root size cannot be parsed from growing environment in that study, it remains possible that differential root size contributed to the observed genotype × environmental interaction for geosmin concentration. To disentangle the effect of root size from that of growing environment on geosmin concentration and, more broadly, to investigate the influence of genotype, growing environment, and fertilizer treatment on geosmin concentration and TDS in table beet, a genotype × environment study is warranted that standardizes root size, compares multiple field environments, and uses mainstream commercial cultivars.
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Genotype mean geosmin concentration and total dissolved solids of 10 individual roots per year of table beet genotypes Bull’s Blood, W357B, Merlin, and Touchstone Gold, averaged over two Wisconsin field sites (Hancock, Madison).
Mean Mg2+ and Ca2+ concentration of ‘Merlin’ table beet leaf tissue grown at two Wisconsin field sites (Hancock, Madison) in 2017. Six plots per site were sampled midseason and at harvest, or 7 and 12 to 13 weeks after planting, respectively.
Total water applied via rain and irrigation, and mean daily temperature from 1 June to 30 Aug. at two Wisconsin field sites (Hancock, Madison) in 2016 and 2017.