Molecular Responses of Nicotiana glutinosa to Infection to Lettuce necrotic yellows virus

Abstract

Lettuce necrotic yellows virus (LNYV), a cytorhabdovirus endemic to Australia and New Zealand, comprises two subgroups: subgroup I (SI) and subgroup II (SII). SI has seemingly become extinct in Australia, potentially outcompeted by SII. This research helps in understanding the host-virus interactions of the model species N. glutinosa with LNYV (Cytorhabdoviridae type species Cytorhabdovirus lactucanecante). This study investigates the molecular and metabolic responses of the model host to infection by each subgroup using one-step reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) and gas chromatography mass spectrometry (GCMS).

Gene expression analysis revealed that both LNYV subgroups induced increased accumulation of AOX1a, NPR3, and RDR1, with LNYV-SII causing weaker responses for AOX1a and RDR1. These results point to a salicylic acid-dependent response by the plant host to infection, implying that LNYV-SII may have evolved mechanisms to dampen the host’s defensive response, potentially contributing to its competitive advantage over SI. In contrast, RDR6 and CPK3 transcript levels were unaffected by LNYV infection. To support these findings, reference genes ACT, EF1α, and SAND were validated for use in LNYV-infected N. glutinosa. EF1α was found to be the most stable, followed by SAND and ACT.

Metabolic profiling using GCMS demonstrated differential levels of amino acids and organic acids in LNYV-SI and LNYV-SII infected plants respectively compared to mock-inoculated plants. This increase in TCA cycle intermediates and activation of the TCA cycle pathway in SII-infected plants implies that these host plants may have low host resistance and hence require more energy to battle viral infection. Compared to SI, the host resistance to LNYV-SII seems to be limited. Higher viral loads are made possible by poor resistance, and this encourages quicker viral multiplication and more efficient vector transmission within plant populations. Common pathways affected by both subgroups included glyoxylate and dicarboxylate metabolism, the citrate cycle (TCA cycle), alanine, aspartate, and glutamate metabolism, and sulphur metabolism. However, SI infection uniquely impacted glutathione metabolism and the metabolism of glycine, histidine, serine, and threonine. Conversely, SII infection uniquely affected arginine biosynthesis and tyrosine metabolism.

These combined molecular and metabolomic analyses provide a greater understanding of LNYV infection in N. glutinosa, highlighting the distinct and overlapping impacts of the two LNYV subgroups. The findings suggest that LNYV-SII’s ability to alter host defence responses and its unique metabolic impacts may explain its rapid spread and competitive displacement of LNYV-SI in Australia. This research represents a significant step towards understanding the molecular process underlying LNYV infection and offers insights into the mechanisms driving the dispersal and dominance of LNYV-SII.