Honeydew collection and chemical analysis

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Method synopsis

Collection of aphid honeydew on aluminum foil substrates followed by LC-MS/MS analysis to quantify sugar and amino acid composition as indicators of plant quality and ant-aphid mutualism benefits.

Synthesized from method descriptions across 2 papers using this protocol.

Procedure as described in the canonical source

Steps below were extracted from the paper that introduces this protocol — Early snowmelt reduces aphid abundance <i>Aphis asclepiadis</i> by creating water stressed host plants <i>Ligusticum porteri</i> and altering interactions with ants (2020), Arthropod-Plant Interactions. Implementations in other papers (listed below) may differ.

1. Collect honeydew samples
   On July 30, placed aluminum foil squares (100 cm²) below extant colonies (N = 50) and left them for 24 hours. Excluded ants from all colonies using barrier. Upon collection, placed foils in individual plastic bags and kept in cooler until frozen in laboratory.
   Quantities: 100 cm² aluminum foil squares, N = 50 colonies sampled, 24 hour collection periodDuration: 24 hour collection period on July 30Conditions: Field conditions, ants excluded during collection
   Equipment: aluminum foil squares, plastic bags, cooler
2. Analyze honeydew composition
   Measured honeydew deposition by counting droplets under stereoscope with image-analysis program. Divided foils into fourths, removed honeydew from one section using ultrapure water, then analyzed composition using high performance liquid chromatography tandem mass spectrometry (LC-MS/MS). Quantified concentrations of six sugars (fructose, glucose, sucrose, trehalose, melezitose and raffinose) and five amino acids (glutamic acid, glutamine, aspartic acid, serine and asparagine).
   Quantities: Foils divided into fourths, 1 section analyzed per foil, 6 sugars + 5 amino acids quantifiedDuration: Laboratory analysis periodConditions: Laboratory conditions, samples kept frozen until analysis
   Equipment: stereoscope (EZ4 HD with LED, Leica Microsystems), image-analysis program (ImageJ, National Institutes of Health, Washington, DC), LC-MS/MS system

Typical Equipment

• aluminum foil squares
• stereoscope
• LC-MS/MS system
• ImageJ

Output Measurements

• honeydew deposition rate
• sugar concentrations
• amino acid concentrations

Papers Using This Protocol (2)

Species Studied With This Protocol

Places Where Used