Epichloë fungal endophyte interactions in perennial ryegrass (Lolium perenne L.) modified to accumulate foliar lipids for increased energy density.

BACKGROUND: Commercial cultivars of perennial ryegrass infected with selected Epichloë fungal endophytes are highly desirable in certain pastures as the resulting mutualistic association has the capacity to confer agronomic benefits (such as invertebrate pest deterrence) largely due to fungal produced secondary metabolites (e.g., alkaloids). In this study, we investigated T2 segregating populations derived from two independent transformation events expressing diacylglycerol acyltransferase (DGAT) and cysteine oleosin (CO) genes designed to increase foliar lipid and biomass accumulation. These populations were either infected with Epichloë festucae var. lolii strain AR1 or Epichloë sp. LpTG-3 strain AR37 to examine relationships between the introduced trait and the endophytic association. Here we report on experiments designed to investigate if expression of the DGAT + CO trait in foliar tissues of perennial ryegrass could negatively impact the grass-endophyte association and vice versa. Both endophyte and plant characters were measured under controlled environment and field conditions. RESULTS: Expected relative increases in total fatty acids of 17-58% accrued as a result of DGAT + CO expression with no significant difference between the endophyte-infected and non-infected progeny. Hyphal growth in association with DGAT + CO expression appeared normal when compared to control plants in a growth chamber. There was no significant difference in mycelial biomass for both strains AR1 and AR37, however, Epichloë-derived alkaloid concentrations were significantly lower on some occasions in the DGAT + CO plants compared to the corresponding null-segregant progenies, although these remained within the reported range for bioactivity. CONCLUSIONS: These results suggest that the mutualistic association formed between perennial ryegrass and selected Epichloë strains does not influence expression of the host DGAT + CO technology, but that endophyte performance may be reduced under some circumstances. Further investigation will now be required to determine the preferred genetic backgrounds for introgression of the DGAT + CO trait in combination with selected endophyte strains, as grass host genetics is a major determinant to the success of the grass-endophyte association in this species.

Changes in Leaf-Level Nitrogen Partitioning and Mesophyll Conductance Deliver Increased Photosynthesis for Lolium perenne Leaves Engineered to Accumulate Lipid Carbon Sinks.

Diacylglycerol acyl-transferase (DGAT) and cysteine oleosin (CO) expression confers a novel carbon sink (of encapsulated lipid droplets) in leaves of Lolium perenne and has been shown to increase photosynthesis and biomass. However, the physiological mechanism by which DGAT + CO increases photosynthesis remains unresolved. To evaluate the relationship between sink strength and photosynthesis, we examined fatty acids (FA), water-soluble carbohydrates (WSC), gas exchange parameters and leaf nitrogen for multiple DGAT + CO lines varying in transgene accumulation. To identify the physiological traits which deliver increased photosynthesis, we assessed two important determinants of photosynthetic efficiency, CO2 conductance from atmosphere to chloroplast, and nitrogen partitioning between different photosynthetic and non-photosynthetic pools. We found that DGAT + CO accumulation increased FA at the expense of WSC in leaves of L. perenne and for those lines with a significant reduction in WSC, we also observed an increase in photosynthesis and photosynthetic nitrogen use efficiency. DGAT + CO L. perenne displayed no change in rubisco content or Vcmax but did exhibit a significant increase in specific leaf area (SLA), stomatal and mesophyll conductance, and leaf nitrogen allocated to photosynthetic electron transport. Collectively, we showed that increased carbon demand via DGAT+CO lipid sink accumulation can induce leaf-level changes in L. perenne which deliver increased rates of photosynthesis and growth. Carbon sinks engineered within photosynthetic cells provide a promising new strategy for increasing photosynthesis and crop productivity.

Red leaf margins indicate increased polygodial content and function as visual signals to reduce herbivory in Pseudowintera colorata.

Red-pigmented leaf margins are common, but their functional significance is unknown. We hypothesized that red leaf margins reduce leaf herbivory by signalling to herbivorous insects the presence of increased chemical defences. Leaves were collected from a natural population of Pseudowintera colorata. Margin size, herbivory damage, anthocyanin content and concentrations of polygodial, a sesquiterpene dialdehyde with antifeedant properties, were quantified. Feeding trials involving larvae of Ctenopseustis obliquana, a generalist herbivore, were conducted on red- and green-margined P. colorata leaves in darkness, or under white, green or red light. Leaves with wider red margins contained higher concentrations of polygodial and anthocyanins, and incurred less natural herbivory. In trials under white light, C. obliquana consumed disproportionately more green- than red-margined leaf laminae. Larvae exhibited no feeding preference when light was manipulated such that leaf colour discrimination was impaired. Red leaf margins provide a reliable and effective visual signal of chemical defence in P. colorata. Ctenopseustis obliquana larvae perceive and respond to the colour of the leaf margins, rather than to olfactory signals. Our study provides direct experimental evidence for aposematic coloration in red leaves.